Dissecting the multiple phenotypes of the yeast Sen1p. Jonathan Sewell Finkel

ISBN: 9781109047769

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NOOKstudy eTextbook

132 pages


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Dissecting the multiple phenotypes of the yeast Sen1p.  by  Jonathan Sewell Finkel

Dissecting the multiple phenotypes of the yeast Sen1p. by Jonathan Sewell Finkel
| NOOKstudy eTextbook | PDF, EPUB, FB2, DjVu, audiobook, mp3, ZIP | 132 pages | ISBN: 9781109047769 | 10.28 Mb

Eukaryotic gene expression is orchestrated by proteins involved in the synthesis of primary transcripts and in co-transcriptional modification of mature RNAs. Genetic analysis has implicated the Saccharomyces cerevisiae superfamily I DNA/RNA helicaseMoreEukaryotic gene expression is orchestrated by proteins involved in the synthesis of primary transcripts and in co-transcriptional modification of mature RNAs.

Genetic analysis has implicated the Saccharomyces cerevisiae superfamily I DNA/RNA helicase Sen1p in gene expression of every major class of RNA. Mutations in SEN1 exhibit multiple nuclear phenotypes in the cell. These include defects in ribosomal RNA processing- efficient intron removal from pre-tRNAs, 3 extended precursors of snoRNAs and snRNAs- and defects in transcription termination/processing of snRNAs, snoRNAs and mRNAs (Steinmetz and Brow, 1996- Ursic et al., 2004- Ursic et al., 1997- Winey and Culbertson, 1988).

Sen1p physically interacts with the C-terminal domain of Rpb1p (largest subunit of RNA polymerase II), with Rnt1p (E. coli RNase III ortholog, dsRNA endonuclease), and with SmD3p (member of the Sm complex required for spliceosome function).-In this thesis I attempted to differentiate the multiple phenotypes associated with Sen1p by examining if the cell directs Sen1p function through specific protein-protein interactions.

Using a variety of methods, mutants were created that resulted in the disruption of the Sen1p-Rnt1p, the Sen1p-SmD3p or the Sen1p-Rpb1p interaction, while maintaining the other established interactions. The effects of these mutants were examined using the transcription and processing of U5 snRNA as an assay.-Using the mutations in SEN1 that specifically impair the interactions of Sen1p with either Rnt1p or Rpb1p, we show that transcription termination and processing are genetically separable events that differentially depend on individual protein-protein interactions.

Specific disruption of the Sen1p-Rnt1p interactions resulted in a defect in the 3 processing of U5 snRNA. Conversely the specific disruption of the Sen1p-Rpb1p interaction caused a transcriptional read-through product of U5 snRNA to accumulate. Our results imply that Sen1p performs separate functions in transcription termination and processing. The approaches used in this thesis are generally applicable to the functional dissection of other complex proteins that engage in multiple protein-protein interactions.



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