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Transcript slippage as a pathway to termination

by
Vadim Molodtsov
Teacher of biology and chemistry, 2003
Russion State Pedagogical University named after A.I. Gertsen

Thesis Advisor: William T. McAllister, Ph.D.

Cell and Molecular Biology Program

Science Center, Room 290

Tuesday, March 2, 2010
12 pm


Abstract

High fidelity of transmission of genetic information during transcription by RNA polymerases (RNAPs) requires that the correct register of the active site is maintained during each cycle of nucleotide incorporation. This lateral stability mainly depends on the Watson-Crick base pairing of the RNA:DNA hybrid. However, it has been observed that when the polymerase encounters homopolymeric tracts in the DNA template the transcript and/or the transcription complex may slip along the template, resulting in incorporation of more or fewer nucleotides than are encoded by the template. This phenomenon, called transcript slippage, may occur during all phases in the transcription cycle including initiation, elongation, and termination. In addition to its role in generating frameshift mutations, transcript slippage has been implicated in promoting the expression of alternative information from the same gene and may play regulatory roles during initiation and termination as well as causing epigenetic changes.

This work was mostly done using bacteriophage T7 RNAP a single subunit enzyme that is capable of performing all stages of transcription. Using specially designed synthetic templates we established the parameters that affect the frequency of transcript slippage by T7 RNAP as well as a few other RNAPs. We also studied the role of transcript slippage in termination at stem-loop termination signals, and demonstrated that termination is promoted in the presence of slippery DNA sequences.

Based on our observations, we propose a new model of transcription termination at stem-loop signals in which the first step in the termination pathway (commitment) is the result of stem-loop assisted slippage, which leads to the formation of a transcriptionally incompetent halted elongation complex that is subsequently followed by complex dissociation.


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