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2007-2008 Gene Regulation Bacterial metabolism Need to respond to changes – have enough of a product, stop production waste of energy stop production.

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Presentation on theme: "2007-2008 Gene Regulation Bacterial metabolism Need to respond to changes – have enough of a product, stop production waste of energy stop production."— Presentation transcript:

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2 2007-2008 Gene Regulation

3 Bacterial metabolism Need to respond to changes – have enough of a product, stop production waste of energy stop production of enzymes for synthesis – new food/energy source, utilize it metabolism, growth, reproduction start production of enzymes for digestion STOP GO

4 Remember Regulating Metabolism? Feedback inhibition – product acts as an allosteric inhibitor – but this is wasteful production of enzymes = inhibition - -

5 Different way to Regulate Metabolism Gene regulation – block transcription of genes for all enzymes in tryptophan pathway = inhibition - - -

6 Gene regulation in bacteria Cells vary amount of specific enzymes by regulating gene transcription – turn genes on or turn genes off turn genes OFF example if enough tryptophan turn genes ON example if new sugar (energy source), like lactose STOP GO

7 Bacteria group genes together Operon – genes grouped together with related functions example: all enzymes in a metabolic pathway – promoter = RNA polymerase binding site single promoter controls transcription of all genes in operon transcribed as one unit & a single mRNA is made – operator = DNA binding site of repressor protein

8 So how can these genes be turned off? Repressor protein – binds to DNA at operator site – blocking RNA polymerase – blocks transcription

9 operatorpromoter Operon model DNATATA RNA polymerase repressor = repressor protein Operon: operator, promoter & genes they control serve as a model for gene regulation gene1gene2gene3gene4 RNA polymerase Repressor protein turns off gene by blocking RNA polymerase binding site. 1234 mRNA enzyme1enzyme2enzyme3enzyme4

10 mRNA enzyme1enzyme2enzyme3enzyme4 operatorpromoter Repressible operon: tryptophan DNATATA RNA polymerase tryptophan repressor repressor protein repressor tryptophan – repressor protein complex Synthesis pathway model When excess tryptophan is present, it binds to tryp repressor protein & triggers repressor to bind to DNA – blocks (represses) transcription gene1gene2gene3gene4 conformational change in repressor protein! 1234 repressor trp RNA polymerase trp

11 Tryptophan operon What happens when tryptophan is present? Don’t need to make tryptophan-building enzymes Tryptophan is allosteric regulator of repressor protein

12 mRNA enzyme1enzyme2enzyme3enzyme4 operatorpromoter Inducible operon: lactose DNATATA RNA polymerase repressor repressor protein repressor lactose – repressor protein complex lactose lac repressor gene1gene2gene3gene4 Digestive pathway model When lactose is present, binds to lac repressor protein & triggers repressor to release DNA – induces transcription RNA polymerase 1234 lac conformational change in repressor protein! lac

13 Lactose operon What happens when lactose is present? Need to make lactose-digesting enzymes Lactose is allosteric regulator of repressor protein

14 Lactose Operon In Presence of Glucose and Lactose, Cell uses glucose. When Glucose is not present and lactose is abundant. Increase in cAMP cAMP binds to CAP Allolactose inactivates lac repressor Transcription on high When Glucose is abundant and lactose is present Low cAMP CAP remains inactive lac Repressor binds to operator

15 Operon summary Repressible operon – Usually in anabolic pathways – end product is in excess Inducible operon – Usually in catabolic pathways, – produce enzymes only when nutrient is available avoids making proteins it does not need

16 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?

17 Points of control in Eukaryotes The control of gene expression at any step in the pathway 1.packing/unpacking DNA 2.transcription 3.mRNA processing 4.mRNA transport 5.translation 6.protein processing 7.protein degradation

18 How do you fit all that DNA into nucleus? – DNA coiling & folding double helix nucleosomes chromatin fiber looped domains chromosome from DNA double helix to condensed chromosome 1. DNA packing

19 Nucleosomes “Beads on a string” – 1 st level of DNA packing – histone proteins 8 protein molecules positively charged amino acids bind tightly to negatively charged DNA DNA packing movie

20 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

21 DNA methylation Methylation of DNA blocks transcription factors – no transcription  genes turned off – attachment of methyl groups (–CH 3 ) to cytosine C = cytosine – nearly permanent inactivation of genes ex. inactivated mammalian X chromosome = Barr body

22 Histone acetylation Acetylation of histones unwinds DNA  loosely wrapped around histones enables transcription genes turned on  attachment of acetyl groups (–COCH 3 ) to histones conformational change in histone proteins transcription factors have easier access to genes

23 2. Transcription initiation Control regions on DNA – promoter nearby control sequence on DNA binding of RNA polymerase & transcription factors – enhancer distant control sequences on DNA binding of activator proteins

24 Model for Enhancer action Enhancer DNA sequences – distant control sequences Activator proteins – bind to enhancer sequence & stimulates transcription Silencer proteins – bind to enhancer sequence & block gene transcription Turning on Gene movie

25 Transcription complex Enhancer Activator Coactivator RNA polymerase II A B F E H TFIID Core promoter and initiation complex Activator Proteins regulatory proteins bind to DNA at distant enhancer sites increase the rate of transcription Coding region T A Enhancer Sites regulatory sites on DNA distant from gene Initiation Complex at Promoter Site binding site of RNA polymerase

26 3. Post-transcriptional control Alternative RNA splicing – variable processing of exons creates a family of proteins

27 4. Regulation of mRNA degradation Life span of mRNA determines amount of protein synthesis – mRNA can last from hours to weeks RNA processing movie

28 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 siRNA

29 Action of siRNA siRNA double-stranded miRNA + siRNA mRNA degraded functionally turns gene off mRNA for translation breakdown enzyme (RISC) dicer enzyme

30 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 Control of translation movie

31 6-7. Protein processing & degradation Protein processing – folding, cleaving, adding sugar groups, targeting for transport Protein degradation – ubiquitin tagging – proteasome degradation Protein processing movie

32 Ubiquitin “Death tag” – mark unwanted proteins with a label – 76 amino acid polypeptide, ubiquitin – labeled proteins are broken down rapidly in "waste disposers" proteasomes

33 Proteasome Protein-degrading “machine” – cell’s waste disposer – breaks down any proteins into 7-9 amino acid fragments cellular recycling play Nobel animation

34 initiation of transcription 1 mRNA splicing 2 mRNA protection 3 initiation of translation 6 mRNA processing 5 1 & 2. transcription - DNA packing - transcription factors 3 & 4. post-transcription - mRNA processing - splicing - 5’ cap & poly-A tail - breakdown by siRNA 5. translation - block start of translation 6 & 7. post-translation - protein processing - protein degradation 7 protein processing & degradation 4 4 Gene Regulation


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