Lecture 11 chapter 12 - Prokaryotic Activators Bacterial...

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12-1 Prokaryotic Activators Bacterial core RNA polymerase is incapable of initiating meaningful transcription RNA polymerase holoenzyme can catalyze only basal level transcription Often insufficient at weak promoters Activators boost basal transcription to higher level via recruitment
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12-2 Eukaryotic Activators Eukaryotic activators also recruit RNA polymerase to promoters indirectly Stimulate binding of general transcription factors and RNA polymerase to a promoter 2 hypotheses for recruitment: General TF cause a stepwise build-up of preinitiation complex General TF and other proteins are already bound to polymerase in an RNA polymerase holoenzyme complex
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12-3 Models for Recruitment of Preinitiation Complex Components in Yeast
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12-4 Recruitment of TFIID e.g. Acidic transcription-activating domain of the herpes virus transcription factor VP16 binds to TFIID in affinity chromatography TFIID is rate-limiting for transcription in some systems TFIID is the primary target of the VP16 transcription-activating domain
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12-5 Recruitment of the Holoenzyme Activation in some yeast promoters appears to function by recruitment of holoenzyme alternative to the stepwise recruitment of individual components of the holoenzyme recruitment of the holoenzyme as a unit is not common
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12-6 Recruitment Model of GAL11P- containing Holoenzyme Dimerization domain of GAL4 binds to mutated GAL11 (GAL11P) in the holoenzyme Interaction does not happen in WT version After dimerization, the holoenzyme, along with TFIID, binds to the promoter, activating the gene
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Activators recruit RNA pol II holoenzyme to promoter -Separable DNA-binding and activation domains -Activation domain binds to one or more subunits of the holoenzyme -Domain-swapping experiments
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12-8 Interaction Among Activators Protein-protein interactions General transcription factors must interact to form the preinitiation complex Activators and general transcription factors interact Activators often interact with one another in activating a gene Individual factors may interact to form a protein dimer facilitating binding to a single DNA enhancer target Specific factors bound to different enhancer sites can collaborate
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12-9 Dimerization Dimerization of an activator increases the affinity between the activator and its DNA target Some activators form homodimers but others function as heterodimers
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12-11 Activators may bind at a Distance Bacterial and eukaryotic enhancers stimulate transcription even when distal to their promoters Four hypotheses for action at a distance Change in topology Sliding Looping Facilitated tracking
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12-12 Hypotheses of Enhancer Action Topology: Sliding: Looping Facilitated tracking
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12-13 3C: Method to detect DNA looping Chromosome conformation capture (3C) to determine if enhancer action requires DNA looping Tests whether two remote DNA regions, such as an enhancer and a promoter, are brought together
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