Gene Expression

• Expression of different set of genes in each cell type

Steps For Regulating Gene Expression

• Transcriptional control is most common

Components For Regulating Transcription

• Short DNA segments of defined sequence• Gene regulatory proteins that bind to a specific sequence

Gene regulatoryprotein

Specific Binding of Gene Regulatory Proteins

• Structural motifs recognize specific DNA sequences• Amino acids interact with outside of bases

Trp Operon

• Multiple genes transcribed as one mRNA molecule• Transcribed from a single promoter

Regulation Of Tryptophan Operon

• On only when tryptophan is absent• Negative regulation by tryptophan repressor• Repressor is active only when tryptophan is bound to

it

Regulation Of Lac Operon

• On only when lactose is present and glucose is absent• Positive regulation by CAP in response to glucose• Negative regulation by lac repressor in response to lactose

Transcriptional Control Region Of Eucaryotic Gene

• Multiple binding sites for gene regulatory proteins• Regulatory sequences known as enhancers can be

thousands of nucleotides from promoter

Eucaryotic Gene Activator Proteins

• Recruitment of RNA polymerase II holoenzyme complex

• Assembly of general transcription factors• Alterations in chromatin structure

Can facilitate:

General function: promote assembly of RNA polymerase II and general transcription factors at the

promoter to allow transcription to begin

Recruitment Of RNA Polymerase II Holoenzyme

• Gene activators attract holoenzyme complex to promoter

Assembly Of General Transcription Factors

• Gene activators promote assembly of some general transcription factors

Alterations In Chromatin Structure

• Recruitment of histone modifying proteins, histone chaperones, and chromatin remodeling complexes

• Can make chromatin more accessible to transcription machinery

Histone Code

• Proteins recognize specific patterns of histone modification• Acetylation promotes activation• Methylation: some residues promote activation some residues promote repression

An example of writing and reading the histone code

Gene Repressor Proteins

TFs in Human Genome1962 estimated (8% of genome)

Gene regulatory proteins, general transcription factors, coactivators, corepressors, chromatin and histone modifiers

Common structural classes of gene regulatory proteins

Zinc finger (762)Homeobox (199)Basic helix-loop-helix (117)Beta-scaffold (87) Basic-leucine zipper (72)Nuclear hormone receptor (49) Forkhead (40) Ets (31)

Examples of TF Classes

Zinc finger Basic helix-loop-helix

• Classes have common motif for DNA binding• Differences within a class determine specificity

Regulating Activity Of Gene Regulatory Proteins

• Modulates pattern of gene expression in response to cell’s environment

DNA Methylation

• Methylation of C at certain CG• Pattern maintained by

maintenance methyl transferases

Effect Of DNA Methylation

• Reinforce inactivation of genes that are not expressed

Genomic Imprinting

• Expression of few genes occurs only from paternal or maternal allele

• Methylation pattern established in germ cells and maintained in offspring

CG IslandsCG

Met

hyl gr

oup

• Many CG lost during vertebrate evolution due to accidental deamination, inefficient repair of methylated CG found at inactive DNA in germ cells

• Promoters of active (often housekeeping) genes in germ cells not methylated, deamination repaired accurately, preserved as CG islands

• CG dinucleotides deficient, preferentially found at promoters of many genes

Epigenetic Mechanisms• Epigenetic inheritance- daughter cells maintain

memory of gene expression pattern of parent cells• Histone modifications, DNA methylation, and

positive feedback loops contribute to epigenetic inheritance

Epigenetic Mechanisms

Histone reader-writers that recognize same histone modification they catalyze

Gene regulatory proteins that activate their own expression

Coordinating Gene Expression

• Decisive event within combinatorial control• Single gene regulatory protein can be decisive,

can control set

Generate Specialized Cell Types

Myogenic helix-loop-helix proteins (MyoD, etc.) and skeletal muscle•Trigger becoming muscle cell•Muscle-specific expression•Coordinately activate muscle genes•Specific for muscle genes

Transcription Attenuation

• Coupled to translation

• Depends upon levels of tryptophan

Attenuation of trp operon:

Alternative Splicing

• Alternative choices for certain splice sites in primary RNA transcript

Regulation Of Alternative Splicing

• Splicing decision controlled by regulatory protein that binds primary RNA transcript

Regulation Of RNA Cleavage In Antibody Genes

• First cleavage site encountered is suboptimal and skipped in unstimulated cells

• Antigen stimulation increases CstF levels to promote cleavage at first site

RNA Editing

A to I editingADAR recognizes RNA structure

C to U editingApoB example

HIV Genome

• Several products through alternative splicing• Some have introns that normally cannot be exported

Regulation Of Nuclear Export Of HIV RNA

• Rev directs export of viral RNAs that contain introns

• Rev levels sufficient to promote export late in infection

Gene Silencing by microRNAs

• miRNAs are dsRNAs processed from hairpin precursors

• miRNA complex binds 3’ UTR of mRNA targets

• Translation repression / mRNA degradation

Translational Repressors

• Bind specific sequences in 5’ or 3’ UTR of RNA

Phosphorylation Of eIF-2

• Activation of specific protein kinases• Reduction of overall protein synthesis by inhibiting

eIF-2B-mediated exchange of GDP→GTP on eIF-2

Regulation Of Translation In Reticulocytes

• Globin translation coordinated with Heme levels• HRI phosphorylates eIF-2• HRI active in absence of heme, inactive in presence of heme

Mechanisms Of mRNA Decay

• Deadenylation-dependent- gradual polyA shortening followed by rapid degradation

• Deadenylation-independent- endonucleolytic removal of polyA

Iron-Mediated Post-Transcriptional Regulation

• Protein that binds to ferritin & transferrin receptor mRNAs• Disassociates from RNA when bound to iron• Blocks translation when bound to 5’ UTR of ferritin mRNA• Stabilizes mRNA when bound to 3’ UTR of transferrin receptor

Aconitase: