BIOL 311 Human Genetics Fall 2006Lecture: Regulation of Gene ExpressionReading: Chap. 10Lecture outline:1. Levels of control2. Histone modification3. RNA polymerase II transcription4. Transcription factors5. Steroid receptors6. Alternate promoters and alternate splicing7. Epigenetic mechanismsLecture:1. Levels of control of gene expressiontranscriptional control- control determines whether or not transcription is initiated- requires promoter of gene- transcription factors bind to promoter and recruit RNA polymerase to initiate transcriptionpost-transcriptional regulation of gene expression- control after pre-mRNA synthesis- can involve control of RNA processing, mRNA transport, translation, mRNA stability, protein processing, protein targeting, protein stabilityepigenetic mechanisms- changes that are heritable, but not due to changes in the order of bases on DNA- main mechanism is DNA methylation. Methylation represses gene expression and determines which allele of a gene is expressed for certain genes (immunoglobulin genes, genes on X-chromosome, etc.)2. Histone modification- histones are basic proteins that help compact DNA- act as generalized repressors of transcription- dense packed chromatin: transcriptionally inactive- to open up the chromatin structure, chemical groups are added to histones to makeconditions more favorable for transcriptionHistone acetylation:1Add acetyl group to epsilon-NH2 group of lysines on histone H3 and H4. Acetylation reduces the positive charge of histone side chains.HATsHistone ---------- acetylated histone -------- HDACs (histone deacetylases)Many other types of modifications of histones occur: methylation, phosphorylation, ubiquitination, ADP-ribosylation.Fig. 10.1 Some histone modifications shown; "histone code"3. RNA polymerase II transcriptionEukaryotes have different RNA polymerases to transcribe different categories of gene.rRNA genes: RNA polymerase I transcribes 18S, 5.8S, 28S rRNA genestRNAs, 5S rRNA, some snRNAs: transcribed by RNA polymerase IIIprotein coding genes and some snRNAs: transcribed by RNA polymerase IIRNA polymerase II12 subunits catalyzes RNA synthesisassociates with about 40 other subunits that are transcription factorsTFIID: TATA binding proteinAlso TFIIB, A, F, E, H4. Transcription factors- basal transcription factors: proteins that assist transcription in many tissues- tissue-specific transcription factors: proteins only present in certain tissues that enhance or repress transcription- bind DNA- may activate transcription directly or bind to other proteins (coactivators) that activate transcription- transcription factors often have conserved domains that mediate protein interactionsDNA binding domains- Zinc fingers, Fig. 10.6 cys2his2 type and cys4 type- Homeodomain (helix-turn-helix) Fig. 10.8- Helix-loop-helix Fig. 10.8Protein interaction domain- Leucine zipper2Activation domains--more difficult to define- Glutamine rich- Proline rich5. Steroid receptorsRespond to small molecules (ligands) in environmentFig. 10.10Ligand ReceptorGlucocorticoids GREstrogen ERThyroid hormone TRVitamin D VDRRetinoic acid RARHave similar DNA binding domains: cys4 zinc fingersLigand binding activates receptorRecognition sites on DNA are similarReceptor--ligand interactions mostly occur in the cytoplasmOther transcription factors mediate environmental responsesChemical signal Transcription factorcAMP CREB/ATFSerum growth factor SRF=serum response factorPhorbol esters AP-1 (dimer of fos/jun)Transcription factors often carry out the final stage of activation, triggered by an environmental signal.Signal + receptor  cell signaling (i.e. kinase cascade) activate transcription factor moves to nucleus to activate gene expression.6. Alternate promoters and alternate splicinghuman genome project- ~35,000 human genes initial predictions were for 80,000-100,000- why the discrepancy?- Single gene can have:multiple transcription start sites (some genes)alternate splicing (many genes): when different selections of exons are used in making different mRNAs from the same geneExample of multiple promoters: - Dystrophin gene 79 exons/2.4 Mb/Xp21 location- Altered in two forms of muscular dystrophy3- Duchenne's MD: severe form, usually due to large deletions in gene- Becker's MD: milder form, usually due to small mutations in gene- At least 7 distinct promoters for dystrophin gene- Active in different tissues- First three promoters when used produce large transcript encoding 427 kDa proteinAlternate splicing and alternate poly(A) addition sites produce many different pre-mRNAs encoding different proteins.Various alternate splicing scenarios: Fig. 10.15Alternate splicing of Wilms tumor protein- Wilms tumor=kidney tumor- WT1 gene altered in some inherited kidney cancers- Single gene encodes 24 isoformso Three possible start codonso RNA editing in exon 6: some RNA bases altered in pre-mRNAo Alternative splicing - Produces isoforms with different functionsDsCam: Drosophila gene with 38,000 possible isoforms7. Epigenetic mechanismsHeritable states which do not depend on DNA sequencesExamples:- DNA methylation- Chromatin remodeling- Histone modificationFig. 10.18Mechanism of DNA methylation- CpG target sequence- Keep the same pattern of methylation as DNA is replicated and passed onWhy is DNA methylated?- Protect from transposons? NO- Regulate gene expression- Generally silences gene expression- Recruits HDAC  leads to deactylation of histone, helping to shut off gene