Quantitative analysis of single molecule tracking in live...
Transcript of Quantitative analysis of single molecule tracking in live...
![Page 1: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/1.jpg)
ProfFredericA.MeunierQueenslandBrainInstitute
TheUniversityofQueenslandAustralia
Quantitativeanalysisofsinglemoleculetrackinginlivecells:fromBrownianmotiontofunction
2018WinterSchoolinMathematicalandComputationalBiologyJuly,2018
Frederic A. Meunier | Queensland Brain InstituteClem Jones Centre for Ageing Dementia Research | Single Molecule Neuroscience laboratory
![Page 2: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/2.jpg)
"Progress in science depends on new techniques, new discoveries and new ideas, probably in that order." Sydney Brenner
![Page 3: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/3.jpg)
- Crashcourseonopticalmicroscopy
- Browsethroughthesinglemoleculelocalizationtechniques
- Singlemoleculeanalysisofthemechanismofneurosecretion(exocytosis)
Thislecture
![Page 5: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/5.jpg)
![Page 6: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/6.jpg)
![Page 7: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/7.jpg)
![Page 8: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/8.jpg)
Superresolutionmicroscopy
![Page 9: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/9.jpg)
FluorescencemicroscopyFluorescence microscopy allows visualisation of the distributions of fluorescently-labelledmoleculeswithinasampleinarelativelynon-invasiveandspecificwayincellsortissues.
Oneofthemajorlimitationistheresolutionlimitsetbythediffractionoflight,whichrestrictstheamountofinformationthatcanbecapturedwithstandardobjectives.Thisisoftenreferredtoasthediffractionbarrier,whichrestrictstheabilityofopticalinstrumentstodistinguishbetweentwoobjectsseparatedbyalateraldistancelessthanapproximatelyhalfthewavelengthoflightusedtoimagethespecimen.Basically:twonearbymolecules=1fluorescentblob…)
Super-resolutiontechniques:inthepastfewyears,anumberofnovelapproacheshavebeenemployedtocircumventthediffractionlimit,includingnear-fieldscanningopticalmicroscopy(NSOM),stimulatedemissiondepletionmicroscopy(STED),stochasticopticalreconstructionmicroscopy(STORM)andstructuredilluminationmicroscopy(SIM).Thesetechniqueshaveallachievedimprovedlateral(x-y)resolutiondowntotensofnanometers,morethananorderofmagnitudebeneaththatimposedbythediffractionlimit,buteachmethodhasauniquesetoflimitations.
![Page 10: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/10.jpg)
Single-moleculelocalizationmicroscopy(SMLM)bypassesthelateralresolutionlimitbyseparatingthefluorescenceemittersintimesothattheyAPPEARoneafteranotherstochasticallyinlocationsthataresufficientlyfarapartfortheiraccuratelocalization.Forthispurpose,photoactivatableorphotoswitchablefluorophoresareused,thusenablingthereconstructionofhigh-resolutionimages.
Typically,SMLMexperimentsareeitherperformedinawide-fieldconfiguration,orintotalinternalreflection(TIR).TheadvantageofTIRfluorescencemicroscopyisthattheexcitationoffluorophoresisrestrictedtoabout100nmalongtheaxialplaneandthusreducestheobservedaxialdepth.Twoconceptsthatrelyonphotoswitchingorphotoactivationaredirectstochasticopticalreconstructionmicroscopy(dSTORM)andphotoactivatedlocalizationmicroscopy(PALM).
Singlemoleculelocalisationtechniques
![Page 11: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/11.jpg)
dSTORMThekeyofSMLMistolimitthenumberoffluorophoresdetectedatagiventimetoallowaccuratelocalizationofsingle-moleculesindividually.InthedSTORMapproach,thisisachievedbyoperatingconventionalsyntheticfluorophores(i.e.commerciallyavailable,forexampleasantibodyconjugates)asphotoswitchesinthepresenceofreducingbuffers.
dSTORMexploitsthefactthatmostfluorophoresarepronetoreductionthatpromotetransitintoalong-lived,non-fluorescentradicalorotherreducedstate(egtripletstate).Thetransitionintothis“dark”stateisgovernedbytheintensityoftheexcitationlight(typicallylaserinduced),andthereducingpotentialofthebufferreagents(typicallythiolssuchasmercaptoethylamine).
Theback-transition(orreturnintothe“bright”(orfluorescent)stateisgovernedbyresidualoxygen,thethermalstabilityoftheoff-stateandtheintensityemittedfluorescence(typicallyblue-shiftedtoabout100nmtotheexcitationlight).ThedSTORMconceptallowsreversiblephotoswitchingofmanycommerciallyavailablefluorophoressuchasareAlexaFluor®647,AlexaFluor®532,ATTO647Netc.
![Page 12: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/12.jpg)
PrincipleofdSTORM.Switchingfluorophoresbetweenadarkandabrightstateenablestemporalseparationofnearbyfluorophores.Fittingofpoint-spreadfunctionsofsingleemittersallowsaccuratepositiondetermination.Super-resolutionimagescanthenbereconstructed.
![Page 13: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/13.jpg)
![Page 14: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/14.jpg)
PALMandsptPALM
Lippincott-Schwartz, J. and Manley, S.
Putting super-resolution fluorescence microscopy to work. Nature Methods 6: 21-23 (2009). A nice overview of the potential benefits and pitfalls of superresolution imaging with emphasis on PALM and related single-molecule techniques. The authors suggest a set of guidelines for presenting images and point out inconsistencies in the current literature.
![Page 15: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/15.jpg)
Daniel Choquet
![Page 16: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/16.jpg)
Nanoscale dynamic organisation of exocytic molecules
Ravi Kasula
Frederic A. Meunier | Queensland Brain InstituteClem Jones Centre for Ageing Dementia Research | Single Molecule Neuroscience laboratory
![Page 17: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/17.jpg)
Li et al., TiBC in press
![Page 18: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/18.jpg)
![Page 19: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/19.jpg)
![Page 20: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/20.jpg)
Martin et al., 2013, J Cell Sci.
Munc18-1Δ317-333 unable to rescue neuroexocytosis in DKD-PC12 cells
![Page 21: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/21.jpg)
Activation Imaging Tracking
20Hzimaging+trackingwithPALM-traceronMetamorph
DKD cells + Munc18-1-mEos
sptPALMRaviKasula
Kasula et al., 2016, Journal of Cell Biology
![Page 22: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/22.jpg)
Munc18-1WTmEos
sptPALM
Kasula et al., 2016, Journal of Cell Biology
![Page 23: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/23.jpg)
![Page 24: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/24.jpg)
![Page 25: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/25.jpg)
Munc18-1WT Munc18-1LM
ControlStimulated
Munc18-1Δ317-333
![Page 26: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/26.jpg)
PALM autocorrelation analysisYeJinChai
![Page 27: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/27.jpg)
Munc18-1WT
![Page 28: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/28.jpg)
Munc18-1Δ317-333
![Page 29: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/29.jpg)
Munc18-1Δ317-333
![Page 30: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/30.jpg)
Munc18-1WT Munc18-1Δ317-333
![Page 31: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/31.jpg)
EffectofMunc18-1onSyntaxin-1nanoscaleorganisation
Kasula et al., 2016, Journal of Cell Biology
![Page 32: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/32.jpg)
Giannoneetal.,BiophysJ2010
Kasula et al., 2016, Journal of Cell Biology
![Page 33: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/33.jpg)
�33Kasula et al., 2016, Journal of Cell Biology
![Page 34: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/34.jpg)
Munc18-1WT + Sx1 Munc18-1Δ317-333 + Sx1
BoNT/ETeTx
Munc18-1WT + Sx1 Munc18-1WT + Sx1
Kasula et al., 2016, Journal of Cell Biology
![Page 35: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/35.jpg)
Munc18-1 domain 3a hinge loop controls the opening of Sx1 and it engagement into the SNARE complex.
Kasula et al., 2016, J Cell Biol.; Bademosi et al., Nature Communications, 2017; Bademosi et al., Jove, 2018, Bademosi et al., Cell Reports, 2018.
- Syntaxin-1 nanoclusters are controlled by NSF and a-SNAP and the PIP2/PIP3 binding domain KARRA of Sx1A (Bademosi et al., Nature Communications, 2017) and… general anaesthetics (Bademosi et al., Cell Reports, 2018).
![Page 36: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/36.jpg)
Opened bunch hypothesis French kiss hypothesis
![Page 37: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/37.jpg)
Morphing potential VAMP2 binding sites of Munc18-1
Residues Identified:
Munc18-1 A297 Munc18-1 T304
VAMP2
We identified two residues that may m e d i a t e M u n c 1 8 - 1 a n d VA M P 2 interaction, closely resembling Vps33 and Vam3 interaction in yeast. (Baker et al. 2015)
Using this we cloned Munc18-1 A297 and T304 residues with Histidine to create steric hindrance and affect its binding with VAMP2
Munc18-1
![Page 38: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/38.jpg)
Increase of Munc18-1 mobility due to its release from nanodomain confinement in DKO-PC12 cells
In agreement with our previous findings in Kasula et al., 2016.
Activity-dependent release of Munc18-1 from nanodomains
![Page 39: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/39.jpg)
Munc18-1 binding to VAMP2 underpins an activity-dependent release of Munc18-1
Mobility of VAMP2 binding deficient Munc18-1 mutants does not increase after stimulation.
Unpublished data
![Page 40: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/40.jpg)
![Page 41: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/41.jpg)
Syn
taxi
n-1A
+Mun
c18-
1WT
Syntaxin-1A-GFP (uPAINT) + Munc18-1WT-mcherry
![Page 42: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/42.jpg)
-5 -4 -3 -2 -1 0 10.00
0.05
0.10
0.15
0.20
0.00 0.05 0.10 0.15 0.200.00
0.05
0.10
0.15
0.5
0.6
0.7
0.8
0.9
1.0
Syn
taxi
n-1A
+Mun
c18-
1A29
7H
MS
D (µ
m2 )
Freq
uenc
y di
strib
utio
n(fr
actio
ns)
Mob
ile fr
actio
n
Time(s) Log(D)
Syntaxin-1A + Munc18-1A297H
![Page 43: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/43.jpg)
-5 -4 -3 -2 -1 0 10.00
0.05
0.10
0.15
0.20
Syn
taxi
n-1A
+Mun
c18-
1T30
4H
0.00 0.05 0.10 0.15 0.200.00
0.05
0.10
0.15
0.6
0.7
0.8
0.9
1.0
MS
D (µ
m2 )
Freq
uenc
y di
strib
utio
n(fr
actio
ns)
Mob
ile fr
actio
n
Time(s) Log(D)
Syntaxin-1A + Munc18-1T304H
![Page 44: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/44.jpg)
Munc18-1WT undergoes an activity-dependent conformational change leading to Sx1 opening and engagement in the SNARE complex.
Conclusions
Munc18-1 domain 3A extended conformation is probably favoured by VAMP2 binding leading to Syntaxin1A opening. Syntaxin-1A opening therefore occurs within the context of vesicular docking (French kiss hypothesis) which may favour proper templating of the SNARE complex during assembly.
![Page 45: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/45.jpg)
Ravi Kasula Adekunle Bademosi Ye Jin Chai Rachel Gormal Sally Martin Andreas Papadopulos Mahdie Mollazade Callista Harper Pranesh Padmanabhan Anmin (Jeff) Jiang Merce Salla-Martret Merja Joensuu Vanessa Lanoue Tristan Wallis Ailisa Blum Chris Small Isabel Morrow Tong Wang Ramon Martinez Marmol
IMB Brett Collins Rob Parton
IINS Bordeaux: Daniel Choquet, Eric Hozy and JB Sibarita UNSW: Yann Gambin, Kat Gaus VIB-KU Louven: Patrik Verstreken Elsa Lawers
Acknowledgements
![Page 46: Quantitative analysis of single molecule tracking in live ...bioinformatics.org.au/winterschool/wp-content/uploads/sites/15/2018/03/Frederic...Morphing potential VAMP2 binding sites](https://reader033.fdocuments.in/reader033/viewer/2022060914/60a8458117b3f15aca790f4f/html5/thumbnails/46.jpg)
PhD and post-doc positions available