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Control of Eukaryotic Genes and Epigenetics AP Biology 2007-2008 The BIG Questions… ■ ■ How are genes turned on & off in eukaryotes? How do cells with the same genes differentiate to perform completely different, specialized functions? AP Biology Evolution of gene regulation ■ Prokaryotes single-celled evolved to grow & divide rapidly must respond quickly to changes in external environment ■ ■ exploit transient resources Gene regulation turn genes on & off rapidly ■ AP Biology flexibility & reversibility adjust levels of enzymes for synthesis & digestion Evolution of gene regulation ■ Eukaryotes multicellular evolved to maintain constant internal conditions while facing changing external conditions ■ homeostasis regulate body as a whole ■ growth & development long term processes ■ specialization turn on & off large number of genes ■ AP Biology must coordinate the body as a whole rather than serve the needs of individual cells Points of control ■ The control of gene expression can occur at any step in the pathway from gene to functional protein packing/unpacking 1. DNA 2. 3. mRNA processing 4. mRNA transport 5. translation 6. protein processing 7. protein degradation AP Biology transcription 1. DNA packing How do you fit all that DNA into nucleus? ■ DNA coiling & folding ■ ■ ■ ■ ■ ■ ■ double helix nucleosomes chromatin fiber looped domains chromosome Link to Animation1 Link to Animation 2 from DNA double helix to AP Biology condensed Nucleosomes ■ 8 histone molecules “Beads on a string” 1st level of DNA packing histone proteins ■ ■ ■ 8 protein molecules positively charged amino acids bind tightly to negatively charged DNA AP Biology DNA packing movie DNA packing as gene control ■ Degree of packing of DNA regulates transcription tightly wrapped around histones ■ ■ no transcription genes turned off heterochromatin darker DNA (H) = tightly packed euchromatin lighter DNA (E) = loosely packed H AP Biology E DNA methylation ■ Methylation of DNA blocks transcription factors no transcription genes turned off attachment of methyl groups (–CH3) to cytosine in ■ ■ C = cytosine nearly permanent inactivation of genes in females(XX) ■ ■ ■ AP Biology ex. inactivated mammalian X chromosome = Barr body why? X VERY LARGE..too much protein if both expressed. Link to Animation (6:05) Methylation a form of Genetic Imprinting (Memory) and silences selected genes. “Pins” represent methyl group additions AP Biology 10 Histone acetylation ■ ■ ■ ■ ■ ■ ■ Acetylation of histones unwinds DNA loosely wrapped around histones enables transcription genes turned on attachment of acetyl groups (–COCH3) to lysine amino acids within the histone protein conformational change in histone proteins transcription factors have easier access to genes Link to Animation Link to Animation AP Biology 2. Transcription initiation ■ Control regions on DNA promoter ■ ■ ■ enhancer ■ ■ ■ AP Biology nearby control sequence on DNA binding of RNA polymerase & transcription factors “base” rate of transcription distant control sequences on DNA binding of activator proteins “enhanced” rate (high level) of transcription Model for Enhancer action ■ Enhancer DNA sequences ■ Activator proteins ■ distant control sequences bind to enhancer sequence & stimulates transcription Silencer proteins bind to enhancer sequence & block gene transcription AP Biology Turning on Gene movie Transcription complex Activator Proteins • regulatory proteins bind to DNA at Enhancer Sites distant enhancer sites • increase the rate of transcription regulatory sites on DNA distant from gene Enhancer Activator Activator Activator Coactivator A E F B TFIID RNA polymerase II H Core promoter and initiation complex Initiation Complex at Promoter Site binding site of RNA polymerase AP Biology 3. Post-transcriptional control ■ Alternative RNA splicing AP Biology variable processing of exons creates a family of proteins 4. Regulation of mRNA degradation ■ Life span of mRNA determines amount of protein synthesis mRNA can last from hours to weeks AP Biology RNA processing movie RNA interference ■ Small interfering RNAs (siRNA) short segments of RNA (21-28 bases) ■ ■ ■ bind to mRNA create sections of double-stranded mRNA “death” tag for mRNA triggers degradation of mRNA cause gene “silencing” ■ ■ ■ ■ post-transcriptional control turns off gene = no protein produced Link to Animation 1 Link to Animation 2 siRNA AP Biology Action of siRNA dicer enzyme mRNA for translation siRNA Text double-stranded miRNA + siRNA breakdown enzyme (RISC) Link to Animation Link to Animation mRNA degraded AP Biology functionally turns gene off RNA interference QuickTime™ and a TIFF (Uncompressed) decompressor are needed to see this picture. QuickTi me™ a nd a TIFF (Uncompre ssed ) decomp resso r are need ed to se e th is p icture. Andrew Fire AP Biology Stanford QuickTi me™ a nd a TIFF (Uncompre ssed ) decomp resso r are need ed to se e th is p icture. Craig Mello U Mass 1990s | 2006 “for their discovery of RNA interference — gene silencing by double-stranded RNA” QuickTime™ and a TIFF (Uncompressed) decompressor are needed to see this picture. 5. Control of translation ■ Block initiation of translation stage regulatory proteins attach to 5' end of mRNA ■ ■ prevent attachment of ribosomal subunits & initiator tRNA block translation of mRNA to protein AP Biology Control of translation movie 6-7. Protein processing & degradation ■ Protein processing ■ folding, cleaving, adding sugar groups, targeting for transport Protein degradation ubiquitin tagging proteasome degradation AP Biology Protein processing movie 1980s | 2004 Ubiquitin ■ “Death tag” mark unwanted proteins with a label 76 amino acid polypeptide, ubiquitin labeled proteins are broken down rapidly in "waste disposers" ■ AP proteasomes Aaron Ciechanover Biology Israel Avram Hershko Israel Irwin Rose UC Riverside Proteasome ■ Protein-degrading “machine” cell’s waste disposer breaks down any proteins into 7-9 amino acid fragments ■ cellular recycling of amino acids ■ Link to Animation ■ Link to Animation AP Biology play Nobel animation 6 7 Gene Regulation protein processing & degradation 1 & 2. transcription - DNA packing - transcription factors (acetylation/methylation) 5 3 & 4. post-transcription - mRNA processing 4 - splicing mRNA - 5’ cap & poly-A tail processing - breakdown by siRNA initiation of translation 5. translation - block start of translation 1 2 initiation of transcription AP Biology mRNA splicing 3 6 & 7. post-translation - protein processing - protein degradation 4 mRNA protection The Ghost in Your Genes Full Documentary on Epigenetics Epigenetics Tutorial from NOVA Epigenetics Tutorial from Cancer Institute Turn your Question Genes on! AP Biology 2007-2008 6 Gene Regulation 7 1 & 2. _________________ - ____________________ - ____________________ 5 4 1 2 3 & 4. _________________ - ____________________ - ____________________ - ____________________ - ____________________ 5. _________________ - ____________________ ____________________ AP Biology 3 4 6 & 7. _________________ - ____________________ - ____________________