Studies of Transcription Termination by Escherichia Coli RNA Polymerase PDF Download
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Author: Karen Marie Arndt
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Category : Escherichia coli
Languages : en
Pages : 362
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Author: Karen Marie Arndt
Publisher:
ISBN:
Category : Escherichia coli
Languages : en
Pages : 362
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Author: Michael John Allen Bellecourt
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Category :
Languages : en
Pages : 0
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Cellular organisms use RNA polymerases (RNAPs) to express genes through the process of transcription. Accurate and timely RNA synthesis requires regulation of all steps of a complex transcription cycle, a regulatory framework involving the careful interplay of interactions between RNAP, the sequences and structure of template DNA and the nascent RNA, and myriad transcription factors that associate with both RNAP and the chromosome. At the end of a transcription unit, after kilobases of transcription, the elongation complex must efficiently release RNA and RNAP from DNA over a 2-3 nucleotide window. In bacteria, there are two programmed termination mechanisms. Intrinsic terminators utilize a GC-rich dyad immediately upstream to a U-rich tract that, together, promotes termination in the absence of additional transcription factors. Rho-dependent termination utilizes the RNA helicase Rho, which binds unstructured RNAs and utilizes its motor activity to extract the nascent RNA. Prior to my studies, the role of nucleic-acid sequence and structure in these mechanisms were well-characterized, but the role of protein conformation changes were largely unknown. To investigate how movements by the RNAP clamp affected intrinsic termination, I disentangled the steps of elongation, commitment, and dissociation, which are conflated in traditional termination measures. I found that clamp movements affected termination efficiency due to effects on elongation, not termination itself. Next, I found that clamp opening is necessary for release of RNAP from DNA, but not release of RNA. I also demonstrated that restricting movements of the trigger loop prior to commitment inhibits intrinsic termination, in agreement with a recently proposed multistate-multipath model of intrinsic termination. My studies also determined how the transcription factor NusG promotes Rho-dependent termination. After binding C-rich RNAs in its secondary site, the Rho ring makes small amino acid rearrangements in its C-terminal domain to induce ring closure. At subpar RNAs that Rho cannot effectively bind, NusG associates near this amino acid switch network, inducing the necessary rearrangements. My studies underscore the importance of protein movements during the termination process, from small-scale Brownian motion to large-scale movements of protein modules, and further demonstrate that RNAP might remain associated with DNA after termination.
Author: David Edward Solow-Cordero
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Category :
Languages : en
Pages : 474
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Author: Laura Magdolna Heisler
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Category :
Languages : en
Pages : 496
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Author:
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Category :
Languages : en
Pages : 375
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Rho is a homohexameric ring-shaped protein that translocates nascent RNA through its central cleft in an ATP-dependent manner, then dissociates RNA polymerase (RNAP) from RNA and template DNA. Despite decades of study, little was known about Rho association with transcription elongation complexes (ECs); sites of Rho termination across the Escherichia coli chromosome; and the effects of the elongation factors NusG, NusA, and the nucleoid structuring protein H-NS on Rho termination. I found that Rho and RNAP have very similar distributions on DNA, suggesting that Rho associates with ECs early and throughout the process of transcription elongation. This association allows Rho to quickly respond to termination signals such as those that are unmasked when transcription and translation become uncoupled. Prior to my studies, the sites of Rho termination in E. coli were mostly unknown. I identified Rho-dependent terminators by examining the distribution of RNAP on DNA in Rho-inhibited cells. I found that Rho terminates highly structured RNAs, such as transfer RNAs (tRNAs) and small RNAs (sRNAs). This finding was surprising, because it was thought that Rho could not associate with structured RNAs. I also found that Rho terminated a small set of novel antisense transcripts that occurred within genes. Upon examining the transcriptome of cell in which Rho was inhibited, I determined that widespread increases in antisense transcription occurred throughout the genome. This finding established that a major function of Rho in vivo is to prevent elongation of antisense transcripts. Finally, I investigated the effects of NusG, NusA, and H-NS on Rho termination. I found that NusG enhances termination at a minority subset of Rho-dependent terminators that are defined by a low C/G ratio at the termination site. In contrast, a large deletion within nusA had no effect on Rho termination in vivo. I also identified a strong overlap between Rho-dependent terminators and H-NS binding sites, as well as genetic interactions between hns and rho. This led me to propose a model in which H-NS enhances Rho termination by slowing RNAP elongation, providing a longer kinetic window for Rho to terminate transcription.
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Author: Patricia L. Tavormina
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Category :
Languages : en
Pages : 340
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Author: Asunción MartÃnez
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Category :
Languages : en
Pages : 466
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Author: Gerald R. Galluppi
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Category : RNA
Languages : en
Pages : 386
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Author: Judith rose Levin
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Category :
Languages : en
Pages : 410
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Author: Ji Lu
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Category :
Languages : en
Pages : 104
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