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Regulation of host RNA polymerase during bacteriophage T7 infection

by
Dhruti Savalia
B.S., 2002
Cook College
Rutgers, the State University

Thesis Advisor: Konstantin Severinov, Ph.D.
Graduate Program in Biochemistry & Molecular Biology

Monday, September 22, 2008
10:00 am


Abstract

T7 is a lytic Escherichia coli (E. coli) phage that encodes a host RNA polymerase (RNAP) inhibitor, gp2. During phage development there is a temporal switch in transcription from expression of early genes, which are transcribed by E. coli RNAP, to expression of middle/late genes, which are transcribed by a T7 RNAP. Due to the ability of gp2 to prevent transcription by E. coli RNAP, the essential role of gp2 could have been the orchestration of this switch in transcription. However, instead of being arrested early in phage development, infection of a wild-type (wt) host by phage without gene 2 is aborted not due to a defect in a switch of transcription from host to viral RNAP but much later at the stage of phage DNA packaging. The primary goal of this work was to elucidate the essential biological role of gp2. The secondary goal of this work was to establish the role of T7-encoded transcription factors in the regulation of host RNAP during phage development.

Our results indicate that during the unproductive infection of a wt host by phage without gene 2, E. coli RNAP continues to transcribe late into the infection from the strong T7 early promoters A1, A2, and A3 which are located at the left end of the genome. However, when the A3 promoter is disabled, unabated early transcription from the other early promoters does not interfere with the success of the infection. Therefore, only continued transcription from A3 is detrimental to infection. Thus, the only essential role of gp2 is to prevent transcription from A3.

T7 is a founding member of a large family of phages which share common genome organization and likely share common mechanisms of gene expression. Bioinformatic analysis of several T7-like phage genomes sequenced and analyzed during the course of this work revealed the presence of the same set of transcriptional regulatory elements and factors as present in T7, indicating that these phages rely on a common transcriptional regulatory strategy during their development.


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