Weaver eoc answers 11-1

Weaver eoc answers 11-1 - Answers to Weaver end of chapter...

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Answers to Weaver end of chapter questions Chapter 11 General Transcription Factors in Eukaryotes 1. List in order the proteins that assemble in vitro to form a class II preinitiation complex. (a) TFIID with help from TFIIA (optional in vitro ) binds to the TATA box forming the DA complex, (b) TFIIB binds next, (c) TFIIF along with RNA polymerase binds to region spanning from -34 to +17. (d) The remaining factors bind in order of TFIIE and TFIIH, forming the DABPolFEH preinitiation complex. 2. Describe and give the results of an experiment that shows that TFIID is the fundamental building block of the class II preinitiation complex. Refer to Fig. 11.1 Reinberg and colleagues performed gel mobility shift assays with different combinations of subunits of the preinitiation complex along with labeled DNA. Experiments without TFIID (lane 13, Fig 11.1a and lane 7, Fig 11.1b) resulted in no complex formation. 3. Describe and give the results of an experiment that shows where TFIID binds. Refer to Fig. 11.2. DNase I based footprinting shows that TFIID protects (binds) the small region just around the TATA box on both the template (Fig 11.2a, DNase I gel, lane labeled D) and non-template (Fig 11.2b, DNases I gel, lane labeled D) strands. 4. Describe and give the results of an experiment that shows that TFIIF and polymerase II bind together, but neither can bind independently to the preinitiation complex. Refer to Fig. 11.1. DNA gel mobility shift assays showed that when TFIIF was added to the DAB complex of transcription factors, there was no detectable difference between the resultant complex and the DAB complex; indicating that TFIIF was not able to bind the DNA (Fig 11.1 compare lanes 2 and 3). However, as increasing concentrations of Pol II was added along with TFIIF, new complexes appeared suggesting that TFIIF and RNA polymerase bind together to the DAB complex (Fig 11.1, lanes 3-7). Similarly, reactions containing decreasing amounts of TFIIF in the presence of Pol II resulted in increasing amounts of just the DAB complex (i.e. lowered higher order complex, Fig 11.1, lanes 8-12). 5. Show the difference between footprints caused by the DAB and DABPolF complexes. What conclusion can you reach based on this difference? Refer to Figs 11.2 and 11.3. The DAB complex forms on the TATA box protecting both the template and nontemplate strands of the DNA, although the DNA at position +10 appears to become hypersensitive to DNase I cleavage. Overall, DAB protects the TATA box region between -17 and -42, whereas DABPolF complex extends this protected region another 34 bases on the nontemplate strand, from position -17 to ~+17. The extended region of protection is consistent with the large size of Pol II and with its binding to one strand of the DNA. 6. Present a hypothesis that explains the fact that substitution of dCs for dTs and dIs for
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Weaver eoc answers 11-1 - Answers to Weaver end of chapter...

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