Brandeis Summer SciFest !

Undergraduate Research Poster Session

Wednesday, August 3, 2011 1:00 – 3:00 pm

Shapiro Science Center Atrium

Poster Titles and Abstracts

Title Understanding the conformational flexibility of human cytochrome P450 3A4

Authors Sophie Kazanis, Jeetayu Biswas and Thomas C. PochapskyAbstract Cytochrome P450 3A4 (CYP3A4) is a microsomal enzyme found in the human liver. It is

responsible for the metabolism of the majority of drugs and xenobiotics that accumulate inthe liver. CYP3A4 can bind and oxidize a wide range of compounds that are structurallydiverse. There is an N-terminal helix found in CYP3A4 that helps bind it to the membrane.Several studies indicate that ligand binding to CYP3A4 is enhanced by interaction with themembrane. Our studies involve examining the conformational flexibility of CYP3A4 usingsolution NMR techniques. Our sample preparations consist of embedding CYP3A4 into ananoscale bilayer ("Nanodisc") which reproduces the native environment accurately.Uniformly labeled 15N, 2H CYP3A4 has been expressed and purified from a bacterialexpression system and embedded into a POPC Nanodisc . Preliminary 15N, 1H TROSY-HSQC experiments have been acquired. In order to further increase the levels ofexpression of CYP3A4 in bacterial systems a fusion protein has been created betweencreated between SUMO (small ubiquitin-related modifier) and CYP3A4 allowing for aneasier purification as well.

Poster # 2011.1Presenter Jeetayu Biswas (Brandeis / Biology, Neuroscience)

Title Atypical Eye Scan Paths of Individuals with Down Syndrome during Face PerceptionTasks

Authors Kevin J. Monk* and Teresa V. Mitchell***Brandeis University, Waltham, MA **E.K. Shriver Center, UMass Medical School,Waltham, MA

Abstract Individuals with Down Syndrome (DS) experience difficulties with face perception tasks,including recognition of emotion and identity. Little is known about the eye gaze behaviorof individuals with DS and whether their difficulties with face perception might be linked toatypical patterns of eye gaze. In the current study, accuracy and eye gaze behaviors ofschool-aged children with DS and typically-developing (TD) controls were recorded asthey performed emotion and identity recognition tasks. Participants viewed a face for threeseconds and then were shown a second face and asked whether the second face was ofthe same emotion (EM task) or the same identity (ID task). While the initial picture wasshown, the scan pattern of the participants was recorded using a special camera.Previous studies in our lab and others show that typical adults direct the majority of theirfixations to the top halves of faces in both EM and ID tasks, with the only deviations to thelower part of the face observed during EM tasks. We observed similar behavior in TDchildren; however this was seen to a lesser extent or not at all in children with DS. Thissuggests that individuals with DS may experience difficulties with face perception becauseof atypical gaze patterns.

Poster # 2011.2Presenter Kevin Monk (Brandeis / Neuroscience)

Title Viabilities of srs2 mutants S890E and S890C are not significantly different than cells witha wild type srs2 plasmid after undergoing a double strand break

Authors Leon Kapulsky, Edwin Antony, Michael Tsabar and James Haber

Abstract Budding yeast, Saccharomyces cerevisiae, undergo mating type switches through a geneconversion event between the MAT locus and either the HMR or HML donor locus. Amating type switch can be initiated by exploiting the galactose-inducible HOendonuclease, which cuts at a site found within the MAT locus of our strains. A cut madeby HO results in a double strand break (DSB), which can then be repaired by geneconversion resulting in a mating type switch. The gene conversion event is mediated byseveral proteins. Two proteins in particular are Rad51 and Srs2. After the DSB, Rad51forms a filament on a piece of single stranded DNA and proceeds to search for homology.In this case of gene conversion, the homology is found on the same chromosome. One ofthe roles of Srs2 is related to its helicase activity. Srs2 is responsible for unwindingnonspecific Rad51 interactions, thus ensuring the fidelity of the repair process. In srs2cells, gene conversion is 20-fold less efficient. This Srs2 activity is mediated by Cdk1,which phosphorylates Srs2, seemingly preventing it from being sumoylated. Biochemicaldata has shown that mutating a phosphorylatable residue of the Srs2 protein results inless unwinding of DNA. In addition, to its role in repair, Srs2 appears to play a role inDNA damage checkpoint recovery. srs2 cells have been shown to be recovery-deficienteven in cells that can repair (as in the case of inter-chromosomal events). To assay theeffects of mutating the 890th residue of Srs2 to either glutamic acid or cysteine, a plasmidwith the corresponding mutation was inserted into srs2 cells and a modified MAT-switching system was used. After DSB, Srs2 wild type plasmid cells had a viability of82.3%; S890E mutants had a viability of 74%; S890C mutants had a viability of 80.5%;and srs2 null had a viability of 4.8%.

Poster # 2011.3Presenter Leon Kapulsky (Brandeis / Biology)

Title Exploring roles of exocytosis and kinesins for protein trafficking associated with sensorycilia in the soil nematode Caenorhabditis elegans

Authors Virginia Ramos, David B. Doroquez and Piali SenguptaDepartment of Biology and Center for Behavioral Genomics, Brandeis University

Abstract The non-motile or primary cilium is a compartment of a cell that functions to facilitateenvironmental sensation. Most of the cells in the mammalian body are ciliated; these ciliahave a variety of functions related to chemosensation, thermosensation, intercellularsignaling, and regulation of fluid flow. Genetic defects that affect the function of cilia leadto a variety of diseases, termed ciliopathies, such as polycystic kidney disease, Bardet-Biedl Syndrome, and retinal degeneration. Since the mechanism underlying ciliadevelopment and maintenance is well-conserved across species from single-cell algae tohumans, we decided to study cilia in a simple model system, Caenorhabditis elegans,where cilia are found at the dendritic tips of sensory neurons. In order to study ciliaformation, maintenance, and protein trafficking we have taken multiple approaches.Specifically, we are studying the role of the exocytosis process with respect to ciliabiology. As null mutations in the exocyst complex genes are lethal for the organism, weare studying the role of these proteins in a tissue-specific manner by knocking-downexocyst function using the method RNA interference (RNAi). We are studying sec-8 andsec-10, two subunits of the exocyst complex, which exhibit a noticeable effect on ciliarygrowth when genetically altered in mammals. We found that transgenic sec-8 C.elegansdisplay irregularities in dye uptake which may be indicative of a defect in cilia morphologyor function. We are currently imaging the cilia to determine if this is the case. Additionally,we would expect to see irregularities in dye-filling uptake and defects in cilia structure insec-10 transgenic worms. We are also investigating whether two kinesin-like proteingenes klp-12 and klp-7 play a role in ciliary trafficking. Our initial approach is to examinethe expression of transcriptional fusion reporters to determine whether klp-12 and klp-7are expressed in ciliated sensory neurons. If we observe sensory neuronal expression, wewill determine where KLP-12 and KLP-7 proteins are localized and the effect of knock-down of gene function via RNAi. If these kinesin like proteins play a role in ciliogenesis weexpect to see dye-fill inconsistencies, as well as defects in cilia morphology. Thesestudies will facilitate our understanding of ciliary biology which may open up avenues fortherapeutic approaches.

Poster # 2011.4Presenter Virginia Ramos (Brandeis / Biology)

Title The Study of mtDNA Mutations During Diabetes in a Model System: Nile Grass Rat

Authors Karina Gaft, Adam Osborne, Fadi Chaabo, KC Hayes and Lawrence WanghAbstract Type 2 diabetes has recently become a worldwide pandemic that affects 26 million people

in the United States alone. Numerous research has linked mitochondrial dysfunction withvarious diseases such as diabetes. A recent study done by the Hayes lab explores theeffects of the diet on a species of rats called Arvicanthis niloticus, the Nile Grass Ratwhich is particularly susceptible to diabetes and contracts symptoms similar to those ofhumans. In this study, the mitochondrial DNA (mtDNA) of Nile Rats will be tracked as itprogresses through the phases of diabetes to confirm any correlations between mutationsin the mtDNA and the onset of diabetes. Using a form of asymmetric Linear-After-The-Exponential PCR (LATE-PCR) which creates abundant single stranded stranded DNAwhich is easily sequenced or probed. By means of the Norwegian Rat sequence as apreliminary data base for the creation of primers, the D-loop region of the Nile Rat hasbeen sequenced. After the COX-1, and Cyt b genes have all been sequenced, primers willbe designed to sequence the whole mitochondrial genome which will be used as a basisfor tracking the mtDNA through the progression of diabetes in the Nile Rat.

Poster # 2011.5Presenter Karina Gaft (Brandeis / Biology)

Title Characterization of the Localization of ObgE, an Essential GTPase Involved inChromosome Partition in Escherichia coli

Authors Jasmine Shackelford, Andrea Cendrowski and Susan T. Lovett

Abstract Chromosome partitioning is a vital stage of the bacterial cell cycle and is inhibited in theabsence of certain proteins. ObgE, an important protein involved in chromosomesegregation, is a highly conserved and essential GTPase in Escherichia coli. Depletion ofObgE results in filamentation and polyploid DNA content. ObgE's cellular localization ispostulated to be important for its function. Visualizing ObgE in vivo is complicated by thefact that fluorescent protein fusions at the N- and C- termini perturb its function andchromosome segregation. The goal of this research was to place GFP-variant tags onObgE, avoiding areas that disrupt its function. Therefore, we engineered three internalObgE-mCherry fusions. Based on previous immunofluorescence data, we predict to seeObgE localize at midcell and hypothesize that its function in chromosome segregation isdirectly related to its cellular position. With a functional fluorescent tag of ObgE, additionalstudies can be conducted to learn more about ObgE and its function in chromosomesegregation and other cell cycle steps.

Poster # 2011.6Presenter Jasmine Shackelford (Spelman College / Biology)

Title Characterization and confirmation of ocrl mutant in Drosophila, a gene implicated in Lowesyndrome

Authors Melissa Lamanna, Sarah Biber and Avital RodalAbstract Lowe syndrome is a rare x-linked disorder that is caused by mutations in the gene ocrl, a

phosphatidyl-inositol phosphatase that is involved in cellular membrane trafficking.However, it is not known how defects in the OCRL enzyme result in the eye, kidney andbrain abnormalities characteristic of Lowe syndrome. We are using the high degree ofhomology between OCRL and the Drosophila melanogaster version of OCRL, dOCRL, tomodel Lowe syndrome in Drosophila's simple and malleable nervous system. Our primaryresearch aim is to generate a null mutation of dOCRL so that we can study how thisaffects cellular trafficking and nervous system function. Twenty-two candidate docrlmutant lines have been generated using one of two different transposases. Thetransposon is expected to excise out of the fly genome near the docrl locus. We hope thatpart or all of docrl will be excised with the transposon, therefore resulting in a null mutationof docrl. We are using site-specific primers and PCR to test if a docrl mutant has beensuccessfully generated. We will use real-time PCR and Western blotting, along with adOCRL specific antibody, to confirm that candidate docrl mutated lines do not express thedOCRL enzyme. A docrl mutant fly line will allow us to better understand the molecularunderpinnings of Lowe syndrome. This is an important step in being able to discoverfuture therapies for the disorder.

Poster # 2011.7Presenter Melissa Lamanna (Indiana University / Biology)

Title The chromatin remodeling protein SWR1 affects gene targeting frequency

Authors Chongsheng Chen, Sarah Dykstra and James HaberAbstract Budding yeast utilizes ATP-dependent chromatin remodeling complexes to enable access

to heterochromatin at the mating-type loci HML and HMR. The chromatin remodelingcomplex SWR1 is known to be a modifier of chromatin structure, acting to exchangehistone H2A with histone H2AZ. The structure of chromatin may play an important role ingene targeting. We generated SWR1 deletion mutants and transformed them with a NAT-marked targeting cassette to determine the effect SWR1 deletion has on the structure ofheterochromatin, and subsequently, the effect SWR1 deletion has on gene targeting. Wefound an effect on gene targeting frequencies when SWR1 was deleted. Deletion of SIR3,a silencing protein, in addition to SWR1, did not seem to significantly affect gene targetingfrequencies.

Poster # 2011.8Presenter Chongsheng Chen (Brandeis / Biology)

Title Identifying Biodiversity Using Nematodes

Authors Patricia Greene, John Rice, Lisa Rice, Arthur H. Reis, Jr. and Lawrence WanghAbstract What is the biodiversity of animals on earth? My research project involves the use of

mitochondrial gene sequences for identification of genera, species, and strains ofnematodes, the largest phylum of multi-cellular animals. A sequences within cytochromeoxidase I gene (COI) is used for identification of variation at the genus and species level,while a sequence within the oxidase III gene (COIII) is used for identification of variationwithin populations of a particular species. These sequences are identical to those used forthe global Barcoding of Life project. DNA sequencing is the standard method of analysisused for such an investigation, but this approach remains expensive and time consuming.Lisa Rice and I have developed a closed-tube LATE-PCR multiplex assay that achieves“Virtual Barcoding” by generating “fluorescent signatures” in four colors. Virtual Barcodingis rapid and relatively inexpensive and can potentially be employed using portabledevices. Our assay will be validated by examining the biodiversity of nematode on theBrandeis campus which includes many diverse micro-environments. Our study will serveas a model for Virtual Barcoding of biodiversity of almost phylum of animals anywhere onearth.

Poster # 2011.9Presenter Patricia Greene (Brandeis / Biology, Anthropology)

Title How Processing Time Affects Memory During Perceptually Difficult Listening

Authors Hannah V Goldberg, Katheryn AQ Cousins and Arthur WingfieldAbstract Previous studies have shown that when words are hard to hear, memory for those words

suffers. This study examines whether this effect can be reduced when more time isallowed for processing. Participants were asked to listen to lists of seven words. Toincrease perceptual difficulty, a single word within a list was acoustically masked withbabble (control lists contained no masked word). Though the babble made the word moredifficult to perceive, the word was still audible. In order to examine the effect ofprocessing time on memory for the words, lists had either a two second or three-secondinter-word interval. Participants were asked to listen to the lists and remember as many ofthe words as they could. When the inter-word interval was 2 seconds long, memory forthe masked word as well as the word prior to the masked word was reduced. We foundthat when the inter-word interval was increased from two seconds to three seconds therewas a recovery of memory performance. A longer processing time allowed masked wordsto be recalled with the same frequency as non-masked words.

Poster # 2011.10Presenter Hannah V Goldberg (Brandeis / Biology, Neuroscience, Philosophy)

Title Single Tube Pathogenic Mycobacterial Detection Assay Using A Fluorescence AnalysisAlgorithm

Authors Skye Fishbein, John Rice, Steve Van Hooser and Lawrence J. WanghAbstract One of the ancient human pathogens, tuberculosis presents a global disease burden,

responsible for over 2 million deaths a year. Public health efforts are hampered by poordetection and inaccurate treatment due to rising drug resistance. Two or three species ofthe infectious Mycobacterium tuberculosis complex (MTBC), that cause humantuberculosis, manifest in unique clinical phenotypes, such as varying drug susceptibility.Thus, identification of the infectious species is necessary for adequate treatment oftuberculosis. A novel combination of molecular technologies and mathematical algorithmshas been employed for amplification, detection, and analysis of mycobacterial genera andspecies. Using two mycobacterial target genes, analysis produces maximally variant,species-specific fluorescent signatures. Following the LATE-PCR assay, principlecomponent analysis is employed to perform cluster analysis based on the dimensions ofmaximum variance within the PCR data among species. In a single tube, the algorithmseparately identify the most infectious members of the mycobacterial genus. This assaywould enhance the clinician's ability to diagnose and treat a wide variety of tuberculosiscases.

Poster # 2011.11Presenter Skye Fishbein (Brandeis / Biology, Mathematics)

Title The Hydrodynamics of Self-Propelled Particles

Authors Alex Rasmussen, Aparna Baskaran, Arvind Gopinath and Michael HaganAbstract Self-propelled particles are objects that move through a medium by actively exerting

forces on it. Specific realizations of self-propelled particles include bacterial colonies,cytoskeletal filaments and motor proteins and motile cells on substrates and suspensions.Physical interactions such as excluded volume and hydrodynamic interactions amongself-propelled particles lead to macroscopic orientational ordering that is nematic (head-tail symmetric) in character. They also locally develop collective motility and exhibitswarming behavior. In this work, we consider macroscopic dynamical equations for acollection of self-propelled particles - namely for the density, nematic order parameter(measure of orientational ordering) and polarization (magnitude of the collective motility).The aim is to unfold the role of the interplay between orientational ordering and collectivemotility on emergent behavior in these systems.

To this end, we have carried out a linear stability analysis of these equations about boththe homogeneous steady states - isotropic and nematic. We found in our isotropic statethat order and polarization may both become unstable for certain initial densitiesdependent on the speed of rods in the system. Velocity induces fluctuations in order andpolarization with order becoming unstable before polarization. In our nematic state wefound that order may induce polarization in the direction of broken symmetry of the systemat small enough length scales. Our analysis indicated that length scales both parallel andperpendicular to the direction of broken symmetry depended linearly on the square ofvelocity.

To understand the dynamics in the nonlinear regime, we have implemented a numericalsolution of the macroscopic equations. Our preliminary findings suggest that the systemexhibits phase separation behavior, with high density high order regions coexisting withlow density disordered regions. Future work will expand on these preliminary findings tohelp understand the physical mechanisms that underlie emergent behavior in systemssuch as bacterial colonies.

Poster # 2011.12Presenter Alex Rasmussen (Colby College / Mathematics)

Title Retinal Degeneration-Causing Mutations Do Not Affect IMPDH1(546)’s Association withPolyribosomes

Authors Aleze Krumholz, Dharia McGrew, Aimee Butterworth and Lizbeth HedstromAbstract Inosine monophosphate dehydrogenase type 1 (IMPDH1) catalyzes the rate-limiting step

in guanine nucleotide synthesis. IMPDH1 is predominantly expressed in the retina, whereit is present in two forms, IMPDH1(546) and IMPDH1(595). Mutations to the IMPDH1isozyme cause autosomal dominant retinitis pigmentosa (adRP) and are also associatedwith a more severe form of retinal degeneration, Leber Congenital Amaurosis (LCA).IMPDH1 associates with polyribosomes in the retina, suggesting that it may play a role intranslation regulation. The most common RP-causing mutation to IMPDH1, Asp226Asn,disrupts the association of IMPDH1(595) with polyribosomes. Several other mutations toIMPDH1 have been identified in patients with adRP and LCA. If these mutations have asimilar effect on the retinal isoforms of IMPDH1, then they may play a role in thepathological mechanism of IMPDH1-mediated retinal disease. The effects of thesemutations on polyribosome association of retinal isoform IMPDH1(546) have been testedand preliminary results suggest that these mutations do not appear to have a significanteffect on the association. Since Asp226Asn was previously shown to cause a defect inpolyribosome association in the minor (595) variant, in the future the effects of these othermutations on the IMPDH1(595) retinal isoform will be investigated.

Poster # 2011.13Presenter Aleze Krumholz (Brandeis / Biology)

Title Prolonged monocular deprivation induces oscillating network activity in primary visualcortex

Authors Alyssa Tyas, Mary Lambo and Gina TurrigianoAbstract During the developmental critical period visual deprivation induces significant changes in

visual responses. It has been shown that monocular deprivation (MD) induces an initialdecrease in responsiveness of the cortical neurons of the deprived eye, followed by aslower increase in responsiveness to the non-deprived eye. Although it is known thatseveral plasticity mechanisms contribute to these changes it is unknown how they affectoverall cortical network activity. To address this issue, we measured the expression of anactivity-dependent, early growth protein, Egr1 in the monocular region of primary visualcortex (V1M) after 1, 3, and 6 days of MD. We sutured one eye of p21 or p22 Long-Evansrats and performed profusions after 1, 3, or 6 days, then stained for NeuN and Egr1proteins. Quantification of Egr1 expression after 1day of MD in V1M showed a decrease inthe deprived hemisphere relative to the control hemisphere (deprived: 83.85% ±3.51% ofmean grey-value), an increase after 3 days of MD (deprived: 126.47% ± 3.77% of meangrey-value) and another decrease after 6 days of MD(deprived:45.18% ± 2.50% of meangrey-value). The findings were similar after normalizing the raw intense density byperimeter and area (perimeter 1d: 82.17% ± 5.76%; area 1d: 83.85% ± 3.51%:perimeter 3d: 131.58% ± 6.32%; area 3d: 126.47% ± 3.77%: perimeter 6d: 40.69% ±3.18%; area 6d: 45.16% ± 2.50%). This data provides evidence that prolonged MDinduces an initial decrease followed by a delayed increase and a further delayed decreasein overall network activity, suggesting that reduced sensory drive leads to a slowoscillation in overall network activity. The delayed increase in activity could reflect ahomeostatic process in response to loss of sensory input, followed by a correctivedecrease due to an overshoot of the baseline.

Poster # 2011.14Presenter Alyssa Tyas (University of Massachusetts Amerst / Biology)

Title Purification of an ~800 kDa complex from S. cerevisiae

Authors Tianyun Wu, Clarisse van der Feltz, Nikolaus Grigorieff and Daniel Pomeranz KrummelAbstract The long-term goal of my project is to prepare intact yeast (S. cerevisiae) U1 snRNP using

the tandem affinity purification (TAP) method for biochemical characterization andstructural study by electron microscopy. Structure of the yeast U1 snRNP will reveal thecomplex interaction network of its RNA subunit and numerous proteins. Further, it willprovide insight into the role of U1 snRNP in splicing - in particular, a critical role it has inrecognition of precursor-mRNAs. This summer, troubleshooting of the TAP protocol andassessment by electron microscopy have been performed.

Poster # 2011.15Presenter Tianyun Wu (Brandeis / Biochemistry)

Title Construction of Tandem Fusion mCherry Cassette for Better Visualization of YeastProteins

Authors Emily Arriola, Riko Hatakeyama and Satoshi Yoshida

Abstract Green fluorescent proteins (GFPs) are an important tool in biological research for studyinglocalization and dynamics of a molecule. Development of fluorescent protein variants withdifferent emission colors has enabled us to use multi-color visualization. To make betteruse of mCherry (a red fluorescent protein) in yeast cell biology, we first designed andconstructed a plasmid which can be conveniently used for tagging any proteins of interestin yeast with mCherry. Although mCherry is one of the brightest RFPs (red fluorescentproteins), it is still half that of GFP and is difficult to visualize in the cells. To overcome thisproblem, we have also constructed a tandemly fused mCherry cassette for brighterimaging. To verify the usefulness of the mCherry cassette and the brightness of themCherry fusion, we have introduced mCherry tag to several yeast proteins. Developmentof the tandem Cherry cassette will allow mCherry to be more easily imaged and will allowease of tagging with a one-step PCR.

Poster # 2011.16Presenter Emily Arriola (Texas A&M University-Corpus Christi / Biology)

Title Structure-function analysis of the Nervous Wreck F-BAR domain and TAP-tagging theendogenous protein

Authors Kelly Miao, Agata Becalska and Avital RodalAbstract Nervous Wreck (Nwk) is a member of the F-BAR family of proteins that drive lipid

tubulation during endocytosis and vesicle trafficking. In Drosophila, nwk mutants causetemperature sensitive paralysis and disrupt coordination of growth signaling betweenneurons and larval muscles at the neuromuscular junction (NMJ). In order to gain a betterunderstanding of the mechanisms by which Nwk drives lipid tubulation during endocytosis,we have engineered a series of Nwk mutants and truncations based on structurepredictions and protein homology. The mutants are designed to disrupt highly conservedmotifs in Nwk. The truncations attempt to create F-BAR domain constructs that aresoluble in order to facilitate in vitro experiments. Currently we have been able todetermine that all three of the Nwk F-BAR fragments are not soluble in E. coli lysate. Inaddition in order to determine additional Nwk interactors in vivo, we are engineering a C-terminal tandem affinity purification tag (TAP) tag on endogenous Nwk by homologousrecombination.

Poster # 2011.17Presenter Kelly Miao (Brandeis / Biology)

Title Effects of Synthetic Multivalent Ligands Displaying Glutamine and Tryptophan onHuntingtin Protein Aggregation

Authors Stephanie M. Armanious and Jason K. PontrelloAbstract The biological role of protein aggregation in Huntington's Disease, a genetic brain disorder

that causes the gradual deterioration and loss of neurons in certain regions of the brain,has yet to be determined. To explore the consequences of protein aggregation, we willsynthesize a series of synthetic multivalent ligands displaying the amino acids glutamineand tryptophan. We hypothesize that glutamine, which has been shown to target thepolyQ repeat in the huntingtin protein, will be able to bind multiple copies of the protein,triggering aggregation. In addition, we believe that targeting the polyproline helix of thehuntingtin protein using the indole sidechain of tryptophan, may enhance the binding ofour synthetic multivalent ligands to the huntingtin protein targets. By varying the averagepolymer length, we anticipate developing compounds that function as either inducers orinhibitors of huntingtin protein aggregation. Thus far, we have synthesized a generalpolymer designed to display two ligands in a block arrangement. One ligand will beattached through an amide bond while the other ligand will be attached through afunctional group explored in this work. The block copolymer was synthesized from an N-hydroxysuccinimide (NHS) ester-substituted norbornene (for an attachment of an amine-bearing ligand) and a Boc-protected amine substituted norbornene (for attachment of asecond ligand). For the future, we are planning to attach glutamine and tryptophan to ourgeneral block copolymer and determine the biological effectiveness of synthesizedcompounds using peptide aggregation assays as well as microscopy techniques.

Poster # 2011.18Presenter Stephanie Armanious (Brandeis / Biology, HSSP)

Title The Role of Sema5A in Mammalian CNS Synaptogenesis

Authors Julie Wertz, Jessie St. Martin and Suzanne ParadisDepartment of Biology, Brandeis University, Waltham, MA

Abstract The semaphorin family is a family of molecules that are involved in axonal guidance aswell as several other processes such as cell migration and angiogenesis. Members of theclass 3 and class 4 semaphorin families, and Sema5B have previously been shown to beinvolved in synaptogenesis. Semaphorin 5A (Sema5A) is an integral membranesemaphorin that can act as either an attractive or a repulsive guidance cue with respect todeveloping axons. Sema5A is downregulated in people with autism and a humangenome-wide association study has revealed that SEMA5A is an autism susceptibilitylocus. One possible explanation for the cognitive effects of autism spectrum disorders isaberrant synapse formation and/or function. Thus, in order to determine whether and howSema5A is involved in glutamatergic synaptogenesis, we designed shRNAs targetingSema5A. We assessed their efficacy by co-transfecting HEK 293T cells with the shRNAsand Sema5A cDNA, and monitored Sema5A levels by Western blotting. Next, we knockeddown Sema5A expression in neurons using the shRNA constructs and assayed excitatorysynapse density by immunostaining for synaptic markers and subsequent confocalimaging. We assessed dendritic branching by Sholl analysis. Preliminary results indicatethat Sema5A knockdown decreases glutamatergic synapse density and increasesdendritic branching. Further research is necessary to confirm these findings.

Poster # 2011.19Presenter Julie Wertz (Brandeis / Biochemistry)

Title Application of Multivalent Displays on Metalloprotease-Dependent Cleavage ofSemaphorin 4D

Authors Frank Anthony Scangarello, Aram Raissi, Jason K. Pontrello and Suzanne ParadisAbstract Semaphorin 4D (Sema4D) is a transmembrane protein expressed in the brain and has

been shown to regulate the development and maturation of GABAergic synapses via anunknown mechanism. Preliminary studies in the Paradis laboratory have shown thatSema4D is cleaved in HEK293T cells and neurons with accumulation of an intracellular,C-terminal fragment. Using a Western Blot assay to monitor appearance of the Sema4DC-terminal cleavage product, we have demonstrated that Sema4D cleavage ismetalloprotease-dependent in HEK293T cells using the metalloprotease activator, PMA,and broad spectrum metalloprotease inhibitor, GM6001. We are currently developingmore selective and higher affinity metalloprotease inhibitors to target Sema4D cleavage inneurons. To this end, we are in the process of synthesizing a broad spectrummetalloprotease inhibitor with the anticipation of using a multivalent presentation toenhance inhibitor avidity (binding) and selectivity by increasing the local concentration ofinhibitor. To optimize multivalent inhibitor effectiveness, we willsystematically vary the valency (polymer length) and inhibitor density, gauging the effectsof these structural variations using our Western Blot assay. We will then apply the mosteffective inhibitor to rat hippocampal neurons to test the role of metalloproteases onSema4D cleavage. Furthermore, due to our modular synthetic route using the RingOpening Metathesis Polymerization (ROMP)-derived polymer scaffold, we anticipatemodification of this inhibitor structure to allow for protein identification. Incorporation of aphotoaffinity label onto the polymer scaffold will allow covalent crosslinking to the targetbound enzyme. Additionally, incorporation of a biotin tag will allow isolation of thecomplex, followed by use of mass spectrometry to identify the unknown metalloproteasethat may regulate Sema4D cleavage and ultimately, Sema4D-dependent inhibitorysynaptic formation.

Poster # 2011.20Presenter Frank Anthony Scangarello (Brandeis / Biochemistry, Biology, Chemistry)

Title VPS35: Purification of a Challenging Protein

Authors Sophie Travis, Ce Feng Liu, Dagmar Ringe and Gregory PetskoAbstract VPS35 forms the core of the retromer complex, a highly conserved heterotrimer involved

in endosome-to-Golgi transport. Unfortunately, VPS35 appears to be highly unstable invitro, and unfolding of VPS35 is thought to be a contributing factor in Alzheimer's disease.Because of its challenging physical properties, VPS35 has not yet been fullycharacterized. This summer, I have cloned and purified an N-terminal fragment of VPS35that appears to have improved solubility and stability over the full-length protein. In future,I intend to screen crystallization conditions of this fragment in hopes of determining astructure.

Poster # 2011.21Presenter Sophie Travis (Brandeis / Biochemistry, Chemical Biology)

Title Cryo-electron tomography and 3D-modeling indicate that the hyperstable ribbon is locatedat the partition between the A- and B-tubule of doublet microtubules in sea urchin spermflagella

Authors Peter Warren1, Xiaofeng Fu1,2, Daniela Nicastro1 and Richard Linck3

1 Department of Biology, Brandeis University2 Howard Hughes Medical Insitutue3 Department of Genetics, Cell Biology and Development, University of Minnesota,

Abstract Doublet microtubules that form the cytoskeleton axoneme are essential for the assemblyand stability of flagella and cilia. Genetic defects that affect the assembly of motile andsensory cilia are known to cause human diseases, such as polycystic kidney disease,situs inversus and infertility. Previous studies conducted on the doublet microtubules ofsea urchin sperm flagella shown that Sarkosyl detergent treatment dissolves the doubletswith the exception of a specialized set of 3-4 protofilaments of the A-tubule. Theseprotofilaments, named the "Sarkosyl-insoluble ribbon", are composed of several proteins,including the conserved, structural protein known as tektin. The exact location of these"Sarkosyl-insoluble ribbons" has been a controversy for the past 40 years. To localize theribbon and tektin protein, we are using cryo-electron tomography and image processingtools to reconstruct the 3D structure of thermal fractionated A-tubules, i.e. partiallydissolved doublet microtubules leaving the hyperstable protofilaments. We determinedthat the hyperstable ribbons extend from protofilaments A11-A13 which forms the partitionbetween the A-tubule and B-Tubule of doublet microtubules. In the future, we will confirmby antibody labeling if this partition ribbon contains tektin, which would verify that thepartition is the ‚"Sarkosyl-insoluble ribbon". If tektin is not present in the partition, thisindicates that there is more than one stable ribbon in doublet microtubules. We believe theincreased understanding of the ciliary and flageullar structure of the axonemes, willprovide greater understanding of these organelles in health and disease.

Poster # 2011.22Presenter Peter Warren (Brandeis / Biology)

Title Nucleic acid binding of Drosophila Inosine Monophosphate Dehydrogenase: a study ofstructure and function

Authors James Chin, Yuliya Mints and Lizbeth HedstromAbstract Transcription is a fundamental cellular process required for cell growth and proliferation,

regulated by a region of deoxyribonucleic (DNA) acid called the promoter. The associationof inosine-5!-monophosphate dehydrogenase (IMPDH), a metabolic enzyme involved inthe synthesis of nucleic acid precursors, with the promoter is unexpected. The associationof IMPDH at the promoter has been attributed to a subdomain of IMPDH in vitro. I proposethat this subdomain is required for IMPDH association with nucleic acids in vivo. Thishypothesis will be tested by investigating the associations of IMPDH mutants with thepromoter in Drosophila S2 cells. This project will identify the structural determinants ofIMPDH promoter association.

Poster # 2011.23Presenter James Chin (Brandeis / Biochemistry)

Title Monitoring the Capacity of Short-Term Memory

Authors Nicole Amichetti, Alison White, Raymond Stanley and Arthur WingfieldAbstract This study aims to examine individuals' ability to monitor their own short-term memory

capacity. Specifically, we examine whether individuals are aware of their own memorycapacity and whether this will be affected by perceptual effort. It has been suggested thatsuccessful comprehension of degraded speech would draw resources that mightotherwise be available for memory. Consequently, when resources are expended forhearing there are fewer resources left for memory. We hypothesize that in difficultlistening conditions an individual's memory ability will be compromised. In twoexperiments we examined how more effortful listening can affects recall and the ability toencode information in memory. We found that listeners hearing words with a softerpresentation level recalled fewer items even though the words could be correctlyidentified. In addition, they consistently overestimated their memory capacity relative tolisteners who were presented with easily audible words. This type of difficult listening andits effect on memory are of concern to those with hearing loss. In the future, we willspecifically examine the older adult population because of the prevalence of hearing lossin the community. It is evident that research surrounding age-related disorders isnecessary because, among older adults, hearing loss is the third most prevalent chronicmedical condition.

Poster # 2011.24Presenter Alison White (Brandeis / Psychology)

Title Identification of adult pheromone receptors in C. elegans using a candidate geneapproach

Authors Michelle Wang, Scott J. Neal and Piali SenguptaAbstract The world is a dynamic and unpredictable place; as a result, animals have developed

remarkable capabilities to cope with their ever-changing surroundings. For example, infavorable environments, the nematode Caenorhabditis elegans develops rapidly toreproductive maturity, but in adverse environments, these animals arrest at the stressresistant dauer diapause stage. Dauer formation is primarily regulated by foodabundance, temperature, and the concentration of dauer pheromone, which is comprisedof a blend of small molecules. This pheromone serves as the basis for C. elegansintraspecific communication and has been shown to not only induce dauer formation butalso elicit robust adult behaviors. Although several pheromone receptors mediating dauerformation have recently been described, the chemoreceptor(s) mediating adultpheromone responses remain unknown. Three pairs of sensory neurons, ADL, ASK, andAWB, are known to be central to the adult pheromone response in C. elegans, and weseek to identify which receptors are expressed in these cells. We have devised a list ofpotential receptors and have attached green fluorescent protein reporters to each of theirpromoters in order to determine whether or not these candidates are expressed in theadult pheromone-sensing neurons. We will then clone the cDNAs of the receptors meetingthis criterion and express them specifically in the ASH neurons. Previous work hasindicated that ASH neurons do not respond to pheromones but demonstrate specificcalcium signals when established pheromone receptors are mis-expressed in them.Therefore, the observation of specific calcium events in the ASH neurons will providestrong support for the conclusion that our candidate receptor is a genuine pheromonereceptor. An understanding of how pheromone signals are transduced in C. elegans willenhance our knowledge of animal communication.

Poster # 2011.25Presenter Michelle Wang (Brandeis / Biology, Economics)

Title The Effect of Sympathetic Innervation on Cardiomocyte Maturation and Cell Size inNeonatal Rat Pups

Authors Chika Okafor, Rebecca Kreipke and Susan BirrenAbstract In the United States, cardiovascular disease is the leading cause of death, accounting for

25% of deaths in 2007. In 2006, nearly 2300 Americans died daily of cardiovasculardisease. Cardiovascular illness is also the most prevalent congenital ailment. There is anoticeable difference in innervation of healthy heart tissue and heart tissue in a diseasedstate, suggesting the effects of the sympathetic nervous system is of great importance.Prior to birth, cardiomyocytes undergo hyperplasic growth. Shortly after birth,cardiomyocytes withdraw from the cell cycle and continue to grow via hypertrophyCardiomyocytes grow via two pathways of hypertrophy: pathological and physiological.Physiological hypertrophy occurs under normal conditions of bodily stress such aspregnancy and exercise. Pathological hypertrophy occurs after heart illnesses.Understanding the mechanisms underlying the maturation of cardiomyocytes beforeexiting the cell cycle may lead to a form of chemotherapy to revert pathologicallyhypertrophic cells back to the cell cycle. In this work, we aim to determine thedevelopmental characteristics of neuronal and cardiomyocyte maturation in a coculturesystem. Previous work has shown that sympathetic neurons halt the increase ofhypertrophic growth, acting, at least in part, via noradrenergic signaling. We then askedwhether these sympathetic neurons play a corresponding role in the control of thewithdrawal of cardiomyocytes from the cell cycle. Cardiomyocytes were grown in thepresence or absence of neurons and some were treated with the beta-adrenergicantagonist propranalol for 2 to 8 days. We showed that cardiomyocytes withdraw from thecell cycle along a similar time frame as that seen in vivo. We also demonstrated thatsympathetic neurons may play a role in regulating the dynamics of cardiomyocyte cellcycle withdrawal and that there may a role for beta-adrenergic signaling. This suggeststhat the sympathetic nervous system is a key regulator of the developmental switch fromcardiomyocyte growth via hyperplasia to hypertrophy.

Poster # 2011.26Presenter Chika Okafor (Florida A&M / Biology)

Title The role of membrane traffic in Drosophila TDP-43 models of ALS neurodegeneration.

Authors Christina Luo, Emily Messelaar and Avital Rodal.Abstract Amyotrophic Lateral Sclerosis (ALS) is a prevalent neurodegenerative disease caused by

loss of upper and lower motor neurons, resulting in death within 1-5 years of diagnosis.Unfortunately, no effective therapies exist for the disease. Recent genetic studies havelinked mutations in TAR DNA binding protein-43 (TDP-43) to heritable forms of ALS, andaggregates of TDP-43 are found in sporadic ALS. TDP-43 is an RNA-binding protein withhundreds of targets, and it is not known how TDP-43 defects lead to neurodegeneration.Studies in Drosophila melanogaster have shown that overexpressing human TDP-43causes neurodegenerative phenotypes, including shortened life span and defective eyedevelopment. Drosophila is therefore a good model system to dissect the cellularmechanisms of TDP-43-induced neurodegeneration. One hypothesis is that alteredgrowth and survival signaling by trophic receptors leads to neurodegeneration. Trophicsignaling is also regulated by membrane traffic, and manipulation of membrane traffic maybe a method to counteract the effects of TDP-43.We set out to reproduce the reported neurodegenerative phenotype in a transgenic line ofDrosophila expressing the human TDP-43 protein (TDP-43L2) by measuring life span andclimbing ability. We also did preliminary crosses of the TDP-43L2 line to two endocyticmutant lines, Vps35 and Snx16, in order to determine whether these mutationsaugmented or ameliorated the neurodegenerative phenotype seen with TDP-43L2 alone.We found that the TDP-43L2 flies had a significantly reduced life span and a fasterdeterioration in their ability to climb up vials compared to control flies. These resultsrecapitulate the earlier study and support the conclusion that overexpression of TDP-43leads to neurodegenerative phenotypes. We also show that the phenotypic differencebetween the overexpression TDP-43 baseline and the endocytic mutants overexpressingTDP-43 is subtle. We are currently conducting full-scale Vps35 and Snx16 crosses on thelarge scale of the crosses done for the overexpression TDP-43L2 line alone. A largersample size will provide a more statistically significant result and enable us to make amore confident conclusion on the effects of these endocytic mutants on the TDP-43pathogenic phenotype. In future studies, we will do these assays on other endocyticmutant lines crossed to overexpression TDP-43 in order to further characterize the cellularbasis of neurodegeneration resulting from pathological changes in TDP-43.

Poster # 2011.27Presenter Emily Messelaar (Brandeis / Neuroscience)

Title A Systems Level Look at Phototaxis in Chlamydomonas reinhardtii

Authors Melissa Donahue, Shani Aharon, Arvind Gopinath, Dongshin Kim and Azadeh SamadaniAbstract Chlamydomonas reinhardtii (Chlamy) is a microscopic unicellular alga that uses

phototaxis to position itself in an optimal environment for photosynthesis. Previousexperiments have examined the response of the flagella of a single trapped Chlamy tolight of different wavelengths and intensities. We propose a complimentary set ofexperiments to examine the phototactic response of large numbers of swimming Chlamy.The overarching goal is to identify the systems level features which enable the Chlamy todetect and to follow light sources. To do this, the effect of inter-Chlamy interactions on thephototaxis of a single cell through hydrodynamic effects and the characteristics ofswimming trajectories such as run length and turning time to perceived light gradients andlight intensities must be understood. After testing various materials, it was determinedthat PEG caused the least number of Chlamy to adhere to the surface. Chlamy wereviewed on PEG coated glass slides using phase microscopy, and the videos are analyzedusing MatLab to track position, swimming speed, phototactic index, and diffusivity. It wasconcluded that in the absence of a light gradient, the phototactic index ofChlamydomonas is close to 0. Future directions include using fluorescent latex beads toquantify the activity of Chlamydomonas under various intensities and creating a lightgradient under which we can observe Chlamy.

Poster # 2011.28Presenter Melissa Donahue (University of Massachusetts Amherst / Chemical Engineering)

Title Metal Complexes Containing a Tridentate Pincer-Type Ligand Featuring a CentralPhosphenium Donor

Authors Zhequan Xu and Christine M. ThomasAbstract In recent years, environmental problems caused by excess release of carbon dioxide and

methane due to the consumption of fossil fuels have stimulated research on a drastic shiftin the world's energy production from fossil fuels to clean and sustainable energy. Myresearch focuses on the synthesis of transition metal complexes featuring a non-innocentligand with an N-heterocyclic phosphenium cation; I seek to synthesize a family of highlyreactive novel catalysts with the ultimate goal of facilitating the conversion of naturallyabundant small molecules into clean fuels. Synthesis of the NHP pincer ligand backbonewas accomplished via alkylation of o-fluoroaniline with 1,2-dibromoethane to generate adiamine, followed by nucleophilic substitution of the aryl fluorides with KPPh2 to generatea diphosphine. Treatment of diamine with PCl3 in the presence of two equivalents of NEt3

leads to formation of the chlorophosphine precursor. The ligand was coordinated with anumber of transition metals including rhodium, iron, chromium, molybdenum etc. Uponabstraction of halide and reduction of the metal center, a number of metal complexes withthe NHP ligand have been synthesized and characterized. In conclusion, my researchinvolves the coordination chemistry of a variety of transition metals with the N-heterocyclicphosphenium ligand. Future work will focus on the study of their geometry, electronic andcatalytic properties as well as synthesizing other complexes with different metal centers.

Poster # 2011.29Presenter Zhequan Xu (Brandeis / Chemistry)

Title Using Homologous Recombination to form EAG Mutants

Authors Miranda Willacey, Jessica Hutcheson, Peter Bronk and Leslie GriffithAbstract The ether-à-go-go (eag) gene codes for a voltage-gated potassium channel subunit. Loss

of function mutations in Drosophila eag have been shown to affect neuronal excitability,olfaction, associative learning and larval locomotion (4). A possible link between learningand EAG is the fact that the C-terminal tail of EAG binds to Ca2+/calmodulin-dependentkinase II (CaMKII), a protein implicated in learning and memory. In order to study theinteraction between EAG and CaMKII, we are generating eag mutant flies with mutationsthat inhibit the binding of CaMKII to EAG. Other mutations will affect ability of CaMKII tophosphorylate EAG. The final mutants will then be analyzed with an associative learningparadigm to study the effects of altering the interaction of EAG and CaMKII on associativelearning.

Poster # 2011.30Presenter Miranda Willacey (University of North Texas / Biochemistry)

Title Parkinson's Disease: Why the Gender Bias?

Authors Nana Owusu, Brandon Meiseles, Jared Auclair, Kristin Boggio and Jeffrey Agar

Abstract Studies have shown that women have a smaller (2-3 fold less) risk of developingParkinson's Disease (PD) and Dementia with Lewy Bodies (DLB) than men. Others havehypothesized that protection from PD results from gender-specific differences inproteolytic processing of toxic proteins. Brain tissue extracts from both males and femaleswere therefore separated first based on size and subsequently analyzed by massspectrometry. In this way, differences in proteolytic processing of alpha-synuclein, aprotein implicated in PD/DLB, and proteins in general, could be determined.

Denaturing gel electrophoresis is the tool frequently used to separate proteins from cellextracts, tissues, and other biological components. When proteins of low-molecular weight(i.e. in the order of 2.5 to 50 kDa) are to be separated and analyzed, however, theresolution of standard SDS-PAGE technique is insufficient. Ideally, tricine gels are used toseparate these low-mass proteins. A tricine gel system was therefore used to separatesoluble protein extracts from human brains affected with a PD-like disease, DLB. Peptidesamples obtained after tryptic digest of the PD samples will be analyzed by Matrix-Assisted Laser Desorption/Ionization-Time of Flight (MALDI-TOF) and MALDI-TOF/TOF.To our knowledge, this is the first time an in-gel digestion has been conducted inconjunction with tricine gels to analyze protein samples using Bottom-up proteomics.

A band of approximately 5 kDa was observed in female PD samples but not males. In-geldigestion of this band is currently being performed and we expect to use Bottom-Upproteomics to identify what this protein is exactly.

Poster # 2011.31Presenter Nana Owusu (Brandeis / Biology)

Title Actin Fluctuations in a Nematic Background

Authors Carlos Perez, Mark Zakhary, Andrew Ward and Zvonimir DogicAbstract A theory by M. Dennison of Utrecht University states that the ends of the polymer, actin,

will deviate from the nematic director to a greater degree then that of the bulk, in asuspension of the fd virus. In order to either support or refute this theory we suspendedactin in a magnetically aligned nematic background at several different concentrations andmeasured it deviations from the nematic director. The direction of the nematic director wasfound using polscope imagining, which measures the retardance produced by the nematicfd background. This combined with fluorescence imaging allowed us to measure thedeviations of the actin filaments from the nematic director and determine whether bends inthe actin filament are pre-existing or are caused by the nematic background. We expectour results will demonstrate that the average deviation of the ends of the actin filament willbe greater then that of the bulk, thus supporting the theory proposed by M. Dennisonconcerning polymers in a nematic background of the fd virus.

Poster # 2011.32Presenter Carlos Perez (Brandeis / Physics)

Title Subcloning of GFP-labeled GABAAR subunits and their expression in neurons

Authors MacKenzie Dwyer1, Ilona Chudotvorova2 and Suzanne Paradis2

(1) Earlham College, Richmond, IN.(2) Biology Department, Brandeis University

Abstract Proper synaptic formation is critical during the development and maturation of themammalian central nervous system (CNS). It includes the precise assembly of proteincomplexes at both the presynaptic and the postsynaptic compartments. GABAergicsynapses are the major type of inhibitory synapse in the mammalian hippocampus.Despite recent progress made in understanding the formation of excitatory synapses earlyin development, formation of GABAergic synapses remains poorly elucidated. In order tobetter understand the mechanisms of inhibitory synapse formation early in development,we subcloned GFP into plasmids expressing the "1, " 2, and #3 subunits of GABAARs.Next, we transfected each vector into HEK293T cells and cultured rat hippocampalneurons and observed expression of the GFP-tagged GABAAR subunits. Using theseconstructs, we will perform time-lapse fluorescence microscopy of transfected neuronsduring early stages of synapse development and during a period of robustsynaptogenesis. These studies will help illuminate the development of inhibitory synapses.

Poster # 2011.33Presenter MacKenzie Dwyer (Earlham College / Neuroscience, Japanese Studies)

Title The Role Of Mph1 And Sgs1 Helicases In Modulating Gene Targeting Efficiency inSaccharomyces cerevisiae

Authors Rosmel Hernandez, Ranjith Anand and James E. HaberAbstract Gene targeting is a technique for DNA manipulation that replaces the target DNA

sequence with the DNA sequence of interest (donor) by making use of shared homologiesbetween them. The process typically includes a method to deliver DNA into the cells.Subsequent to its internalization, the donor is subject to processing by various enzymespresent inside the cell nucleus that includes DNA resection, strand invasion/assimilationand integration. Thus, many cellular factors can affect gene-targeting efficiency. Usingbudding yeast Saccahromyces cerevisiae as the model organism, we wish to determinethe role of Sgs1 and Mph1 helicases in modulating gene targeting efficiency. Sgs1 is a3’-5’ helicase that is known to be involved in DNA resection. Mph1, also a 3’-5’ helicase,has been shown to be involved in the creation and disruption of strand invasionintermediates. These helicases have also been shown to negatively affect a homologydependent repair pathway called Break Induced Replication (BIR) in which only one endof the broken DNA shares homology with the donor. Gene targeting also involves "one-ended" events at both sides of the targeting fragment. In order to address the roles of theabove helicases in gene targeting, DNA a construct coding for the URA3 gene was madethat will be used to replace the endogenous THR4 gene located in chromosome III. Theefficiency of gene targeting will be initially determined in the yeast mutants lacking Sgs1(sgs1$) and Mph1 (mph1$) helicases by selecting for the cells that have become Ura+and Thr–. Experiments will also be conducted on yeast strains lacking both the helicases(sgs1$mph1$). An mph1 mutant was made by replacing the endogenous MPH1 genewith the nourseothricin resistance marker (NAT). Sgs1 mutant will be made by replacingSGS1 gene with Kanamycin resistance marker (KAN). Double mutants will be made eitherby simultaneous replacements with the respective markers or by consecutive replacementof sgs1 in an mph1$ strain. Isogenous wild-type (WT) haploid yeast strain that is eitherMATalpha or MATa will be used as controls. Measurements of targeting efficiencies inthese strains will help us better understand the process of gene targeting.

Poster # 2011.34Presenter Rosmel Hernandez (Brandeis / Biology)

Title From proton to chloride pump: the D93T mutation in deltarhodopsin

Authors Amy R. Eisenberg, Kristine Mackin and Douglas L. TheobaldAbstract Rhodopsins comprise an important group of membrane transport proteins with

homologous structures and functions. Bacteriorhodopsin (bR) uses the key residue Asp-85 (D85) and a Schiff base to pump protons. Based on sequence alignment, both bR anddeltarhodopsin (dR), another proton pump, have this conserved Asp, whereashalorhodopsin (hR), a chloride pump, has a threonine in the homologous position.Previously, bR was converted to a chloride transporter by mutating D85 to threonine(D85T). Here, we provide preliminary evidence that dR, which is expressed in active formin E. coli, is converted from a proton to a chloride pump by incorporating the mutation atthe corresponding position: D93T.

Poster # 2011.35Presenter Amy Eisenberg (Brandeis / Biology)

Title Organizing Circadian Rhythms through Firing of Drosophila Clock Neurons

Authors Isadora Cerullo, Michael Rosbash and Fang GuoAbstract In flies and mammals, circadian rhythms are controlled by endogenous pacemakers that

run on an approximately 24-hour period but also respond and synchronize toenvironmental cues like light and temperature. This robust yet flexible circadianmechanism allows organisms to be active during the most favorable times of the day.Light-mediated phase shifting - producing delays in the early morning and advances in thelate night - is considered cell-autonomous in Drosophila, triggered by CRYPTOCHROME(CRY)-mediated photon capture and then TIMELESS (TIM) degradation within most if notall of the ca. 150 circadian neurons of the adult brain. However, previous results from ourlab reveal that a light pulse in the early night leads to variable TIM degradation withindifferent clock neurons. This suggests an important role for circadian neuroncommunication. Our goal was therefore to determine how neuronal activity is integratedinto the circadian rhythm mechanisms of Drosophila. More specifically, can the firing ofcircadian neurons entrain the phase of circadian protein oscillations and produce circadianactivity/sleep patterns? This was tested by briefly activating a subset of pacemakerneurons. This firing induces a phase-shift during the subjective day and also impacts thecircadian phase of flies without functional CRY. Furthermore, temporal firing of particularclock neurons under constant light conditions produces at least two cycles of self-sustained oscillations in these normally arrhythmic flies. The data, collected by locomotoractivity monitoring, immunohistochemistry, and luciferase assays, indicate that neuronalactivity is an important Zeitgeber (temporal cue) in the fly system and affects the clock atthe molecular and the behavioral level. They also suggest a positive feedback loopbetween neuronal activity and clock protein oscillations.

Poster # 2011.36Presenter Isadora Cerullo (Columbia University / Biology, Human Rights)

Title Regulation of Bnr1 Formin Activity by Smy1

Authors Veronika Kivenson, Julian Eskin and Bruce GoodeAbstract Actin is a highly conserved protein found in eukaryotic cells, and a major component of

the cytoskeleton. Formins are a widely expressed group of large, multi-domain proteinsthat are involved in building actin networks, which are involved in cell motility, shape, anddivision. The yeast S. cerevisiae has two formins: Bnr1 and Bni1, which are involved inassembling actin cables that function as tracks for myosin-dependent intracellulartransport. Bnr1, located at the bud neck, assembles actin cables by nucleating andelongating actin filaments under the regulation of the proteins Smy1 and Bud14, amongothers. Additionally, Bnr1 can bundle filaments in vitro.We are working to determine which surfaces of the Bnr1 FH2 domain are important forbundling (as opposed to nucleation or elongation), and which surfaces are important forinhibition by Smy1 and Bud14. In order to do this, we use site-directed mutagenesis toalter specific Bnr1 residues, and then purify and screen them by using pyrene-actinassembly and bundling assays to check for loss of function. We have purified andquantified Bnr1 and Smy1, and begun to test the effects of mutant proteins on forminfunction. None of the mutants tested to date display dramatic differences from the wildtype protein. In the future, we will continue testing the existing mutants, as well as Smy1mutants.

Poster # 2011.37Presenter Veronika Kivenson (Mount Holyoke College / Biochemistry)

Title Examining differential microRNA expression in cortex during prolonged activity blockade

Authors Sadella Santos, Sean O'Toole and Sacha NelsonAbstract microRNAs (miRNAs) are short, noncoding RNA sequences that play a role in post-

transcriptional regulation. They have been demonstrated to be involved in cell fatedecisions, developmental transitions, as well as cancer and neurological disorders. In thisstudy we decided to examine the potential role of miRNAs in homeostatic plasticity.Homeostatic plasticity manifests itself in several forms, either through scaling the neuron’sresponse to synaptic inputs, adjusting intrinsic properties thus ultimately affecting theneuron’s firing rate, or modulating the release of synaptic vesicles. We hypothesized thatmiRNAs represent a potential mediator of these homeostatic changes. Individual miRNAsare capable of regulating the rate of translation for hundreds of transcripts, severalmiRNAs are known to be regulated in an activity dependent manner, miRNA machinerylocalizes to the synapse and recently one miRNA was demonstrated to participate in apre-synaptic form of homeostatic plasticity in the hippocampus. We asked whether thereis a correlation between levels of miRNA expression and induction of homeostaticchanges within the cortex. We selected a number of miRNA candidates that were knownto be expressed in neurons, were regulated by activity and were demonstrated to affectthe electrophysiological and morphological properties of the synapse. Organotypic slicecultures were treated with Tetrodotoxin (TTX, a drug which blocks voltage gated sodiumchannels therefore decreasing neuronal activity) or kept in control media. The overalllevels of these miRNAs were measured using stem-loop mediated RT-PCR. Expressionlevels were then compared to a reference miRNA (let7a) using the 2-%%CT method. Wefound that the calculated fold changes for these miRNAs were not significant whencomparing slices treated with or without TTX. To make a more confident analysis we willlook into selecting a wider group of miRNAs, increasing the sample size, and increasingthe number of reference genes for our relative comparison.

Poster # 2011.38Presenter Sadella Santos (University of Maryland Baltimore County / Chemical Engineering )

Title Chiral Tweaking of Colloidal Membrane Line Tension

Authors Mir Henglin, Thomas Gibaud and Zvonimir DogicAbstract The filamentous fd viruses serve as a model for rod-like colloids. Aqueous suspensions of

these viruses form classical isotropic and nematic liquid crystal phases. The fd wild-typeand the mutant fd y21m viruses are both chiral and actually form a twisted nematic phase,also known as the cholesteric phase. First, I quantified the chirality of these two virusstrains through measurements of the pitch of their cholesteric phases. A temperature-series measurement of the cholesteric pitch showed that the chirality of the fd wild-type istemperature dependent, while the chirality of the mutant fd y21m is not. Second, adding adepleting polymer agent to aqueous suspensions of these viruses induces a net attractionbetween the viruses, leading to the self-assembly of monolayered colloidal membranes.Though the membranes are mostly composed of aligned viruses, chirality favors twistbetween adjacent viruses. Chirality manifests itself at the edge of the membranes, wherethe viruses form a twisted layer. The membrane edge deforms due to thermal fluctuationsand the line tension of the membrane can be derived from the magnitude of thosedeformations. A temperature series measurement of the membrane line tension of the fdwild-type and mutant fd y21m viruses showed that the line tension of the fd wild typemembranes is temperature dependent, while the line tension of the mutant y21mmembranes is not. The combination of both those experiments, the quantification of thetemperature dependence of chirality and the line tension measurements, unambiguouslydemonstrates the effect of chirality on line tension and rules out any intrinsic temperatureeffects.

Poster # 2011.39Presenter Mir Henglin (Williams College / Physics)

Title Srv2 Enhances Cofilin-Mediated Severing of Actin Filaments

Authors Long Le, Dennis Breitsprecher, Faisal Chaudhry, Kristin Little and Bruce GoodeAbstract Dynamic restructuring of the actin cytoskeleton is a vital component of many cellular

processes, including endocytosis, polarized cell growth and cytokinesis. Rapid actindisassembly allows cells to replenish the available supply of actin monomers for theformation and elongation of new filaments. Srv2/cyclase-associated protein (CAP) from S.cerevisiae belongs to a conserved class of proteins that promote cofilin-dependent actinturnover by recycling cofilin from ADP-actin monomers. Novel findings indicate that the N-terminus of Srv2/CAP harbors an additional activity and directly interacts with cofilin topromote the severing of actin filaments. In order to analyze the effects of Srv2 on cofilin-mediated actin severing, we employed multicolor total internal reflection fluorescence(TIRF) microscopy using purified proteins. This technique is an innovative and powerfultool to analyze actin dynamics at the single-filament level in real time. We expressed,purified and fluorescently labeled several cofilin point mutants, as well as Srv2. MulticolorTIRF was used to observe the interactions between labeled F-actin, labeled cofilin andlabeled Srv2, and to correlate severing events with the localization of cofilin and Srv2 onthe filament sides. Our data show that binding of cofilin to actin filaments directly coincideswith severing events, and that Srv2 enhances the severing efficiency by as much as 5-fold. Moreover, our results suggest that different cysteine labels on Srv2 and cofilininterfere with the cofilin-Srv2 interaction. Thus, more work is needed in order to establishan experimental setup which allows for the direct observation of cofilin/Srv2-interactionsduring filament severing by triple-color TIRF.

Poster # 2011.40Presenter Long Le (Brandeis / Biological-Chemical Engineering)

Title Generation of eag homologous recombination mutants to investigate the non-ionicsignaling of the EAG potassium channel

Authors Jessica Hutcheson, Peter Bronk, Miranda Willacey and Leslie C. GriffithAbstract The Drosophila Ether-à-go-go (EAG) membrane protein is thought to be involved in

memory formation since mutants display memory deficits. In addition to a six-trans-membrane potassium channel subunit region, EAG contains several C-terminal protein-binding motifs including ones for calmodulin (CaM), calcium-calmodulin dependent proteinkinase II (CaMKII), and Drosophila CASK (dCASK). In order to investigate how theseprotein-binding interactions contribute to EAG’s role in learning and memory, we aregenerating several Drosophila eag mutants by homologous recombination of pointmutations into the endogenous locus of the eag gene. More specifically: 1) EAG whichno longer binds CaMKII (R784K/Q785K), 2) mutants in which CaMKII’s phosphorylationtarget on EAG is eliminated (T787A), and 3) EAG mutants that mimic constitutivephosphorylation at this site (T787D) (Sun et al., 2004). A mutant in the CaM-bindingdomain of EAG has already been generated by the same procedure. This mutant hasbeen analyzed by Western blot, and EAG levels have been determined to be similar tothat of wild-type controls, though the effect on CaM-binding is still unknown. Furtheranalysis of all mutants will assess protein binding, electrophysiology, and learningphenotypes.

Poster # 2011.41Presenter Jessica Hutcheson (Brandeis / Neuroscience)

Title Homoallylation of Imines

Authors Stephanie Chun, Isaac Krauss and Wenbo PeiAbstract Although there is a significant body of work on stereoselective allylation of carbonyl

compounds and imines, homoallylation has still not been well investigated. Astereoseletive homoallylation method would add to the arsenal of reactions available fororganic synthesis of complex natural products. Earlier work in our group succeeded inhomoallylating various aldehydes with a cyclopropylmethyl boronate in gooddiasteroseletivity. Our current efforts attempt to expand the substrate scope to includealdehyde-derived imines. As of this writing, none of the imine variations give anystereoisomers of the desired product. Additionally, we are in the process of synthesizinga new boronate with a new methylcyclopropylmethyl group to investigate the range ofsubstituents tolerated on the cyclopropane ring. Future experiments will test a widerrange of imines and increase the substituents on the boronate reagent. Enantiopurealcohols or amines may be able to be synthesized using a new chiral reagent (see Chen &Krauss, Poster 2011.43, for details).

Poster # 2011.42Presenter Stephanie Chun (Brandeis / Chemistry)

Title Synthesis of Enantiopure Boron Reagents for Stereoselective Homoallylation of Carbonyls

Authors Emily Chen and Isaac KraussAbstract Many chemical reactions create structures that are seen in biological products, but the

large majority does not have the appropriate stereochemistry to be useful in biologicalsystems. One such reaction is homoallylation, or the addition of an allyl group with anextra CH2 group to any given chemical structure. The highly desirable bishomoallylalcohol motif created through this process is an important stepping stone to numerousstructures found in bioactive natural products and this novel synthetic route will provideothers with an easier means of creating various medicinally important molecules. Ourgroup has developed a simple and dependable stereoselective homoallylation methodusing cyclopropanated allyl boron reagents. However, only racemic reagents have everbeen used, so the product obtained is a mixture of enantiomers. We are currentlyexploring synthetic routes to produce enantiopure reagents for enantioselectivehomoallylation. To begin, an enantiopure epoxide is converted to a cyclopropyl sulfone,alkylated with a boronic acid pinacol ester and desulfonylated. At present, each stepbefore the desulfonylation has been proven successful, and further experiments willoptimize each step and problem solve through the desulfonylation step.

Poster # 2011.43Presenter Emily Chen (Brandeis / Chemistry)

Title Determining if mutations to IMPDH1(595) have any correlation with polyribosomeassociation and retinal degeneration disease severity

Authors Lael C. Scarborough, Aleze Krumholz and Lizbeth HedstromAbstract Retinitis pigmentosa (RP) is an eye disease that results in the loss of night and peripheral

vision due to damage of the photoreceptor cells. Mutations to IMPDH1(595) cause RPand Leber Congenital Amaurosis (LCA), a more severe version of retinal degeneration,and affect its association with polyribosomes. We tested the hypothesis that disease iscaused by a defect in IMPDH1(595) polyribosome association by transfecting HEK293Tcells with IMPDH1(595) mutant plasmids, isolating polyribosomes with a sucrose cushion,and analyzing the resulting pellet using a western blot. From this experiment, we cansuggest a likely mechanism for these diseases. Specifically, it will allow us to see ifpolyribosome dissociation of IMPDH1(595) is more in mutants causing LCA (N198K andR105W) and less in mutants causing RP (H372P and T116M). Preliminary results suggestthat there may be no correlation between polyribosome association and retinaldegeneration disease severity.

Poster # 2011.44Presenter Lael C. Scarborough (University of Maryland, Baltimore County / Biology)

Title The Dual Role of Magnesium in the Catalytic Cycle of a Kinase

Authors Padraig Niall Murphy, S. Jordan Kerns, Roman Agafonov, Young-Jin Cho and DorotheeKern

Abstract A long-standing and fundamental question in biology is how enzymes are able todramatically decrease the chemical activation barrier and achieve impressiveaccelerations in the rates of chemical reactions. It is now appreciated that conformationalchanges are essential, and in many cases are the rate-limiting steps of the reaction.Adenylate kinase allows us to investigate this complex relationship between the chemicaland physical mechanisms of catalysis due to its large conformational movements ofnucleotide-binding lids around its active site and its ability to catalyze both the forward andreverse reaction. Enzymologists often found Mg2+ to be an essential cofactor inphosphoryl transfer reactions. This forced us to question whether catalysis could occur inthe complete absence of metal. In the absence of Mg2+, the rate of substrate-to-productconversion was quantitatively shown to be 106 fold slower than the Mg2+ catalyzed rate.Through a combination of pre-steady state quench-flow experiments and NMR dynamics,we revealed that phosphoryl transfer becomes the rate limiting step in the absence of Mg2

+. Surprisingly, we also found that Mg2+ accelerates the following step in the catalyticcycle, lid-opening, by three orders of magnitude. We hypothesized that the catalysis ofthe phosphoryl transfer step by Mg2+ is very specific, while the lid-opening acceleration ismainly due to the presence of a divalent metal. Our hypothesis was buttressed byquantifying phosphoryl transfer and lid-opening rates as a function of cationic species. Wefound that magnesium or any divalent cation facilitates lid-opening by weakeningelectrostatic interactions between lid arginines and bound nucleotides. However, Mg2+

increases the rate of phosphoryl transfer by at least 102 over similar divalent cations. In aneffort to elucidate the effect of protonation of the phosphoryl groups on catalytic reactivity,we measured catalytic turnover at low and median pH in a no-metal experiment. We foundthat there was only a small correlation between accelerated rate and median pH. Insummary, our results show the rate of the conformational change, lid-opening/closing, isaccelerated by 103 in the presence of a positive, divalent metal. In contrast, the chemicalstep was found to be much more specific for cations of a certain size and charge.

Poster # 2011.45Presenter Padraig Murphy (Brandeis / Biochemistry, Biophysics)

Title Dense, gravity-driven flow of bidisperse particles in 2-D hopper

Authors Michal Dichter, Bulbul Chakraborty and Shubha TewariAbstract Many systems experience a transition from a fluid-like state to a solid-like state

characterized only by a sudden arrest in dynamics, or "jamming." We show the results ofnumerical simulations of dense, gravity-driven granular flow in a two-dimensional hopperwith a tapered outlet, as in an hourglass. We analyze the velocity at critical areas of thesystem (i) to determine the steadiness or intermittency of flow, and (ii) to search forperiodicity in the flow, both anticipated to be reliant on system geometry, i.e. the particle-diameter-to-outlet-width ratio. An understanding of the causes underlying thephenomenon of jamming in granular flows (aside from being an intrinsically excitingphysics problem) has tremendous technological implications including improved safetyand efficiency in the industrial sector.

Poster # 2011.46Presenter Michal Dichter (Brandeis / Physics)

Title T-cadherin and EphrinB1 act independent of the BMP2 pathway during sympatheticneuron innervation of cardiac myocytes

Authors Angela Chui, Jason J. Baade and Susan BirrenAbstract During development, neurons from the sympathetic nervous system innervate the

maturing heart, which results in neural modulation of cardiac function. It is important tounderstand how sympathetic cardiac innervation is controlled during development sincewhen this is dysregulated it leads to diseases like arrhythmias. Sympathetic neuronalgrowth is inhibited by contact with cardiac myocytes and synapse formation is initiated.One likely candidate molecule to mediate this growth inhibition is T-cadherin (Tcad). Tcadis attached to the cell membrane via a GPI-anchor and is part of the large family ofcadherin adhesion molecules. Tcad has been shown to be sufficient to inhibit the growthof sympathetic neurons, through regulation of neurite growth depending on thedevelopmental state of the cardiac myocyte it contacts. Another possible importantinhibitor of sympathetic neurite growth is EphrinB1, which is also dependent ondevelopmental state. BMP2 (bone morphogenetic protein), a significant factor forembryonic development, promotes sympathetic neurite growth; however, when contact ismade between sympathetic neurons and cardiac myocytes, the BMP2 pathway is alteredand no longer acts as a promoter of neurite growth. Using a co-culture system ofsympathetic neurons, from the superior cervical ganglion (SCG) and cardiac myocytesfrom the heart, both derived from neonatal rats, we tested if either Tcad or EphrinB1functions to alter the BMP2 pathway as part of the interaction between SCG neurons andcardiac myocytes. In order to determine whether either Tcad or EphrinB1 alters the BMP2growth promoting effects as a result of contact, we performed experiments using RNAi toknockdown expression of either Tcad or EphrinB1, in the presence or absence of BMP2.We determined that Tcad does not affect the BMP2 pathway during the interaction ofmature myocytes and sympathetic neurons. With additional experiments, we willdetermine whether EphrinB1 affects the BMP2 pathway during the interaction of youngmyocytes and sympathetic neurons and determine how EphrinB1 affects neurite growthover the area of the cardiac myocytes they innervate.

Poster # 2011.47Presenter Angela Chui (Brandeis / Biology)

Title Surface functionalization as a tool for testing Biomaterial engineering

Authors Katherine Elfer, Rafael A. Cabanas and Seth FradenAbstract The attachment of single stranded DNA and rod-like viruses on different functionalized

surfaces was investigated. Using the same idea as protein microarrays, an attempt ismade to immobilize the Fd virus in order to test their linkage to ssDNA and to the surface.Rod-like viruses are useful for studying soft matter; they are colloidal particles that presentliquid crystalline behavior. Apart from their tendency to orient in the same direction as afunction of concentration, they create different structures when mixed with polymer. Suchbehavior can be modified and controlled by the creation of a DNA network linkingindividual viruses together. Four surfaces were tested with fluorescence microscopy: BSA-biotin, aminopropyl-triethoxysilane, mercaptopropyl-trimethoxysilane (MPTS) and, biotin-PEG-silane. The BSA-biotin surface exhibited uniform attachment of the ssDNA, linkers,and repetitive hybridization of complementary ssDNA when temperatures were cycledabove and below its melting point. The amino-silane surface, when coated with sulfo-SMCC malamide, provides the necessary chemistry for cysteine bonding in a differentvirus strain. The MPTS surface showed cysteine-end bonding with the virus. The biotin-PEG-silane surface initially showed uniform coating, but the coating did not presentequally with every preparation.

Poster # 2011.48Presenter Katherine Elfer (Louisiana Tech University / Nanosystems Engineering)

Title Micro-injection of layer five motor neurons in the cerebral cortex of Amyotrophic LateralSclerosis Mice Brain Tissue: A Mass Spectrometric Study

Authors Emmanuel Obasuyi1, Kristin J. Boggio1, Nathalie Y.R. Agar2 and Jeffrey N. Agar1

1) Brandeis University2) Department of Neurosurgery, Brigham and Women’s Hospital, Harvard Medical School

Abstract Micro-injection and subsequent matrix-assisted laser desorption/ionization time-of-flight(MALDI-TOF) mass spectrometric (MS) analysis is a powerful approach for detectingbiomolecules directly from tissue with high-resolution preservation of spatial information.MALDI-TOF MS is a soft ionization method that ionizes biomolecules such as proteinsand peptides as intact, predominately singly protonated ions, facilitating the determinationof their mass and composition. Micro-injection MALDI-TOF MS provides us with a tool todetect and characterize biomolecules in specific cells still imbedded in tissue sections. Inthis study, we conducted MALDI-TOF analysis of micro-injected layer V motor neurons inthe cerebral cortex of transgenic mice expressing both yellow fluorescent protein (YFP)under control of the Thy1 promoter, and the G93A variant of human Cu/Zn superoxidedismutase, which serves as a model for amyotrophic lateral sclerosis (hSOD1G93A;YFP).Amyotrophic lateral sclerosis (ALS) is a motor neuron disease that is characterized byprogressive degeneration of upper motor neurons in layer V of the cerebral cortex andlower motor neurons in the ventral horn of cervical and lumbar sections of the spinal cord.Brains from mice expressing hSOD1G93A;YFP were dissected at 151 days of age (wheresymptoms are present), and compared to a hSOD1G93A–null, YFP-positive littermatecontrol. Spectra collected from micro-injected neurons in hSOD1G93A;YFP mice showeddecreases in several peak intensities. Both mass peaks at 9979.83m/z and at5444.81m/z showed significant decrease in peak intensity by approximately two-thirds.Serial sections were stained with hematoxylin and eosin (H&E) in order to confirm the lossof motor neurons in layer V of the cortex in hSOD1G93A;YFP mouse. In the H&E stainedserial sections, the loss of motor neurons in layer V of the cortex in hSOD1G93A;YFPmouse can be discerned in comparison to motor neurons in layer V of YFP mouse. Thesefindings could aid in understanding the degeneration of motor neurons in ALS mousemodels based on hSOD1G93A.

Poster # 2011.49Presenter Emmanuel Obasuyi (Brandeis / Neuroscience)

Title Key Players for the Dramatic Rate Acceleration of Phosphoryl Transfer in a Kinase

Authors Lien Phung, Roman Agafonov, Young Jin Cho, Padraig Murphy and Dorothee Kern.Abstract As part of the essential cellular system that maintains energy homeostasis, Adenylate

Kinases (ADK) catalyze the interconversion of 2 ADPs to AMP and ATP. In an effort tounderstand the mechanism by which the kinase interconverts the nucleotides, its crystalstructures and activities were studied. We identified a key residue that takes part in thephosphoryl transfer between the nucleotides. Based on previous experiments on theEscherichia coli ADK (eADK) R156K mutant, it is hypothesized that Arg150 in thehomologue enzyme Aquifex ADK (aADK) would have the similar effect in suppressing theactivity of the enzyme. The residue is thought to be directly involved in the process ofphosphoryl transfer and stabilization of the transition state. The crystal structure of themutant Arg150Lys shows that the lysine does not have contact with the bound nucleotidesand thus can most likely not take part in the catalysis, explaining the observed dramaticdecrease in activity. Kinetic assays by coupled enzymes and pre-steady state quench flowin the presence of magnesium were performed with wild type aADK and the R150Kmutant. The phosphoryl transfer activity of the wild type enzyme is at least 1000 timesfaster than the mutant. Activity experiments without magnesium were done with wild typeaADK and eADK to test the effect of the metal on the enzymes using high-pressure liquidchromatography (HPLC). The phosphoryl transfer rate of the wild type (WT) can only bemeasured in the absence of Mg2+ because the rate with Mg2+ is too fast to be measuredwith current instrumentation. The activity of the enzyme in the presence of Mg2+ is fasterby a magnitude of 106 than when it is free of the metal. These results show that Arg150 inaADK and magnesium are crucial in the catalytic activity of Adenylate Kinases. Based onprevious rate obtained for WT eADK without magnesium of 0.002/sec, the rate of WTaADK is slower at 0.0005/sec. In an experiment with calcium (instead of Mg2+) aADK(1.1476 s-1) was also slower than eADK (12.30 s-1). This surprising result led to thequestioning of the similarity of the active sites of the two homologues. The sequence andstructure of the two enzymes were analyzed. While the direct active site is identical,several satellite residues that either have direct contacts with the nucleotides or the P loopin the enzyme are different. They are possible candidates that constitute the differencebetween the phosporyl transfer rates of the two enzymes. Further investigation with themutant of these residues will be needed to determine the importance of their role in thephosphoryl transfer of the kinase.

Poster # 2011.50Presenter Lien Phung (Brandeis / Biochemistry)

Title Characterization of the Regulatory Behavior of NC2 and Mot1 in TAF-DepletedTranscription

Authors Benjamin D. Longwell1,2, Katie L. Pennington1 and Michael T. Marr II1

1 Department of Biology, Brandeis University, Waltham, Massachusetts 02454, USA2Department of Biology, Washington College, Chestertown, Maryland 21620, USA

Abstract Transcription is a central cellular process necessary for viability of an organism; thereforecharacterizing the roles of factors involved in this process is important for understandingthe molecular nature of life. The eukaryotic transcription factor II D (TFIID) is a keycomponent of the RNA polymerase II (pol II) preinitiation complex. TFIID itself iscomprised of TATA-box binding protein (TBP) and several TBP-associated factors(TAF’s). TBP binding to the TATA box is a necessary step for transcription of TATA-dependent promoters. Although TBP alone is sufficient to facilitate basal-leveltranscription, holo-TFIID is required for activator-dependent transcription at manypromoters However, previous research has shown that when TAFs are depleted,transcription unexpectedly increases at the Drosophila melanogaster metallothionein(Mtn) genes in response to metal-stimulated induction. Additionally, cofactors that affectTBP binding can regulate the rate of transcription. Negative-cofactor 2 (NC2) and modifierof transcription 1 (Mot1) have been shown to inhibit TBP binding to TATA elements. Toassess the possible effect of NC2 and Mot1 on transcription of the MtnA gene, NC2, Mot1,and TAF1 were knocked down in Drosophila melanogaster tissue culture cells. Cells werethen induced with CuSO4 to stimulate transcription of the metallothionein genes.Knockdowns were confirmed by western blot analysis, and the level of transcription wasmeasured by reverse transcription quantitative PCR (RT-qPCR). Knockdowns weresuccessful; western blot analysis showed an approximate 70-90% reduction in all targetgenes. RT-qPCR data support that the depletion of TAFs increases the level oftranscription. The data suggests that NC2# and Mot1 serve primarily as negativeregulators of transcription, and that an increase in transcription when TAF1 is knockeddown is dependent on NC2# and Mot1.

Poster # 2011.51Presenter Ben Longwell (Washington College / Biology)

Title Synthesis, Purification, and Preliminary Characterization of Oxanosine Monophosphate

Authors Philip Braunstein and Lizbeth HedstromAbstract Inosine monophosphate dehydrogenase (IMPDH) is the enzyme that mediates that rate-

limiting step of guanine nucleoside triphosphate biosynthesis. Inhibitors of IMPDH havethe potential to be anti-cancer and anti-parasitic therapeutics. Oxanosine is a naturalnucleoside product isolated from Streptomyces. It has been observed that oxanosinemonophosphate (OMP) is an inhibitor of IMPDH. In this project I will characterize theinhibition of IMPDH by OMP. Here I present synthesis, purification, and preliminarycharacterization of OMP. I will further optimize the purification of OMP and characterizethe kinetics of the inhibitor in the coming year.

Poster # 2011.52Presenter Philip Braunstein (Brandeis / Biochemistry)

Title Characterization of Growth and Cholesterol/Triglyceride Levels in Mice with HumanApolipoprotein A1 Gene

Authors Samuel J Gagne, Fadi Chaabo and KC HayesAbstract Apolipoprotein A1 (APOA1) is the apolipoprotein associated with high-density lipoprotein

(HDL), and is necessary for its function as a cofactor and ligand. Growth curves for knock-in mice with human APOA1 were obtained, and the effect of subtle changes in diets ontotal cholesterol and total triglycerides levels, and LDL/HDL ratio. Mice given both high-fatand high-carbohydrate diets showed an abnormally high level of total cholesterol, as wella human LDL/HDL ratio (mice have a substantially different LDL/HDL ratio than humans).Further trials are necessary to prove the validity of these results, as well as exactcharacterization of lipoprotein levels. The mice's sensitivity to polyunsaturated fats vs.saturated fats is also in the process of being analyzed

Poster # 2011.53Presenter Samuel Gagne (Brandeis / Biology, Neuroscience)

Name Index

Presenter TitlePoster #

Stephanie Armanious Effects of Synthetic Multivalent Ligands Displaying Glutamine andTryptophan on Huntingtin Protein Aggregation

2011.18

Emily Arriola Construction of Tandem Fusion mCherry Cassette for BetterVisualization of Yeast Proteins

2011.16

Jeetayu Biswas Understanding the conformational flexibility of human cytochromeP450 3A4

2011.1

Philip Braunstein Synthesis, Purification, and Preliminary Characterization ofOxanosine Monophosphate

2011.52

Isadora Cerullo Organizing Circadian Rhythms through Firing of Drosophila ClockNeurons

2011.36

Chongsheng Chen The chromatin remodeling protein SWR1 affects gene targetingfrequency

2011.8

Emily Chen Synthesis of Enantiopure Boron Reagents for StereoselectiveHomoallylation of Carbonyls

2011.43

James Chin Nucleic acid binding of Drosophila Inosine MonophosphateDehydrogenase: a study of structure and function

2011.23

Angela Chui T-cadherin and EphrinB1 act independent of the BMP2 pathwayduring sympathetic neuron innervation of cardiac myocytes

2011.47

Stephanie Chun Homoallylation of Imines2011.42

Michal Dichter Dense, gravity-driven flow of bidisperse particles in 2-D hopper2011.46

Melissa Donahue A Systems Level Look at Phototaxis in Chlamydomonas reinhardtii2011.28

MacKenzie Dwyer Subcloning of GFP-labeled GABAAR subunits and their expressionin neurons

2011.33

Amy Eisenberg From proton to chloride pump: the D93T mutation in deltarhodopsin2011.35

Katherine Elfer Surface functionalization as a tool for testing Biomaterialengineering

2011.48

Skye Fishbein Single Tube Pathogenic Mycobacterial Detection Assay Using AFluorescence Analysis Algorithm

2011.11

Karina Gaft The Study of mtDNA Mutations During Diabetes in a ModelSystem: Nile Grass Rat

2011.5

Samuel Gagne Characterization of Growth and Cholesterol/Triglyceride Levels inMice with Human Apolipoprotein A1 Gene

2011.53

Hannah V Goldberg How Processing Time Affects Memory During Perceptually DifficultListening

2011.10

Patricia Greene Identifying Biodiversity Using Nematodes2011.9

Mir Henglin Chiral Tweaking of Colloidal Membrane Line Tension2011.39

Presenter TitlePoster #

Rosmel Hernandez The Role Of Mph1 And Sgs1 Helicases In Modulating GeneTargeting Efficiency in Saccharomyces cerevisiae

2011.34

Jessica Hutcheson Generation of eag homologous recombination mutants toinvestigate the non-ionic signaling of the EAG potassium channel

2011.41

Leon Kapulsky Viabilities of srs2 mutants S890E and S890C are not significantlydifferent than cells with a wild type srs2 plasmid after undergoing adouble strand break

2011.3

Veronika Kivenson Regulation of Bnr1 Formin Activity by Smy12011.37

Aleze Krumholz Retinal Degeneration-Causing Mutations Do Not Affect IMPDH1(546)’s Association with Polyribosomes

2011.13

Melissa Lamanna Characterization and confirmation of ocrl mutant in Drosophila, agene implicated in Lowe syndrome

2011.7

Long Le Srv2 Enhances Cofilin-Mediated Severing of Actin Filaments2011.40

Ben Longwell Characterization of the Regulatory Behavior of NC2 and Mot1 inTAF-Depleted Transcription

2011.51

Emily Messelaar The role of membrane traffic in Drosophila TDP-43 models of ALSneurodegeneration.

2011.27

Kelly Miao Structure-function analysis of the Nervous Wreck F-BAR domainand TAP-tagging the endogenous protein

2011.17

Kevin Monk Atypical Eye Scan Paths of Individuals with Down Syndrome duringFace Perception Tasks

2011.2

Padraig Murphy The Dual Role of Magnesium in the Catalytic Cycle of a Kinase2011.45

Emmanuel Obasuyi Micro-injection of layer five motor neurons in the cerebral cortex ofAmyotrophic Lateral Sclerosis Mice Brain Tissue: A MassSpectrometric Study

2011.49

Chika Okafor The Effect of Sympathetic Innervation on Cardiomocyte Maturationand Cell Size in Neonatal Rat Pups

2011.26

Nana Owusu Parkinson's Disease: Why the Gender Bias?2011.31

Carlos Perez Actin Fluctuations in a Nematic Background2011.32

Lien Phung Key Players for the Dramatic Rate Acceleration of PhosphorylTransfer in a Kinase

2011.50

Virginia Ramos Exploring roles of exocytosis and kinesins for protein traffickingassociated with sensory cilia in the soil nematode Caenorhabditiselegans

2011.4

Alex Rasmussen The Hydrodynamics of Self-Propelled Particles2011.12

Sadella Santos Examining differential microRNA expression in cortex duringprolonged activity blockade

2011.38

Presenter TitlePoster #

Frank AnthonyScangarello

Application of Multivalent Displays on Metalloprotease-DependentCleavage of Semaphorin 4D

2011.20

Lael C. Scarborough Determining if mutations to IMPDH1(595) have any correlation withpolyribosome association and retinal degeneration disease severity

2011.44

Jasmine Shackelford Characterization of the Localization of ObgE, an Essential GTPaseInvolved in Chromosome Partition in Escherichia coli

2011.6

Sophie Travis VPS35: Purification of a Challenging Protein2011.21

Alyssa Tyas Prolonged monocular deprivation induces oscillating networkactivity in primary visual cortex

2011.14

Michelle Wang Identification of adult pheromone receptors in C. elegans using acandidate gene approach

2011.25

Peter Warren Cryo-electron tomography and 3D-modeling indicate that thehyperstable ribbon is located at the partition between the A- and B-tubule of doublet microtubules in sea urchin sperm flagella

2011.22

Julie Wertz The Role of Sema5A in Mammalian CNS Synaptogenesis2011.19

Alison White Monitoring the Capacity of Short-Term Memory2011.24

Miranda Willacey Using Homologous Recombination to form EAG Mutants2011.30

Tianyun Wu Purification of an ~800 kDa complex from S. cerevisiae2011.15

Zhequan Xu Metal Complexes Containing a Tridentate Pincer-Type LigandFeaturing a Central Phosphenium Donor

2011.29

Sponsorship for Brandeis Summer SciFest provided by:

Department of Biology, Brandeis University

Division of Science, Brandeis University

Research Experiences for Undergraduates Program, "Cell and Molecular Visualization at Brandeis University"

National Science Foundation

Research Experiences for Undergraduates Program, Materials Research Science and Engineering Center

National Science Foundation !