Transcription_probs

Transcription_probs - MCB121 C S Gasser MCB121 Problem Set...

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MCB121, C. S. Gasser MCB121 Problem Set: Transcription Machinery 1. For each of the following statements referring to aspects of eukaryotic gene expression circle which types of genes (type I, II, or III) to which the statement applies. Circle ALL correct answers for full credit. I II III Transcription complex includes TATA-binding protein (TBP). I II III Transcription usually requires TFIID. I II III RNA product can be translated into protein. I II III RNA product can be a component of a ribosome. 2. The diagram below illustrates the results of a gel shift experiment. Two DNA fragments, DNA 1 and DNA 2, are suspected of including binding sites for two newly discovered general transcription factors (factor X and factor Y). The fragments were radioactively labeled an used in the gel shift assay with purified factors X and Y. The information above each lane indicates which components were added to the reaction run in that lane. Migration is from top to bottom. Results for the last lane (at right) are not given. A. For each of the following circle “T” if the statement is true, “F” if the statement in false, and “N” if the truth of the statement cannot be determined from the information given. T F N Factor X can bind to DNA 1 by itself. T F N Factors X and Y can form a heterodimer in the absence of DNA. T F N Factor X only associates with DNA 1 in the presence of Factor Y. T F N The arrow at right indicates the band corresponding to factor Y bound to DNA 1. T F N The two bands indicated by the star at right are identical in their composition. T F N Free DNA 1 migrates to the same position as free DNA 2. Factor X Factor Y DNA 1 DNA 2 2 X 2 Y 2 1 X 1 Y 1 X Y 1 X Y 1 2 X Y 2 *
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MCB121, C. S. Gasser B. (5) In the last lane on the gel above, draw the bands you would expect to see from a reaction containing all four components (factors X and Y, and DNAs 1 and 2). 3. The C-terminal domain (CTD) of the large subunit of RNA polymerase II consists of multiple repeats of the consensus sequence TyrSerProThrSerProSer. This region appears to have important roles in polymerase function. This region can be phosphorylated by CTD-kinase (CTD-K) activity and can be dephosphorylated by CTD phosphatase (CTD- P). You have recently discovered specific inhibitors of CTD-K and CTD-P which you call I K , and I P , respectively. In vitro transcription reactions are carried out on a standard DNA template in reactions including all normally required transcription factors (with initially unphosphorylated Pol II), and including CTD-K and CTD-P. Four reactions are performed, one with no inhibitor, one with I K , one with I P , and one with both inhibitors present (inhibitors are added before adding the DNA template in each case). A. (12) What would the relative levels of transcript be in the four reactions (which reaction would produce the most, least etc. transcript)? Explain briefly.
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