The role of Clp1 and Pcf11 in transcription and pre-mRNA 3’-end processing
Eukaryotic transcripts require a number of complex cotranscriptional modifications and processing events before translation to protein. Clp1 and Pcf11 are subunits of cleavage factor IA (CFIA), an essential component of the Saccharomyces cerevisiae pre-mRNA 3’-end processing machinery. The crystal s...
Main Author: | |
---|---|
Published: |
University College London (University of London)
2012
|
Subjects: | |
Online Access: | http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.568289 |
id |
ndltd-bl.uk-oai-ethos.bl.uk-568289 |
---|---|
record_format |
oai_dc |
spelling |
ndltd-bl.uk-oai-ethos.bl.uk-5682892016-11-18T03:18:16ZThe role of Clp1 and Pcf11 in transcription and pre-mRNA 3’-end processingHedden, J. J.2012Eukaryotic transcripts require a number of complex cotranscriptional modifications and processing events before translation to protein. Clp1 and Pcf11 are subunits of cleavage factor IA (CFIA), an essential component of the Saccharomyces cerevisiae pre-mRNA 3’-end processing machinery. The crystal structure of a Clp1-Pcf11 complex was determined previously and revealed the binding of ATP to a highly-conserved P-loop motif and a tight Pcf11-Clp1 interaction facilitated by a number of highly-conserved Pcf11 residues. Nonetheless, the biological function of both Clp1-ATP binding and the Pcf11-Clp1 interaction was not well understood. The work in this thesis combines an in vitro and in vivo investigation of the Clp-ATP and Clp-Pcf11 interactions in an effort to understand the function of these factors in transcription and pre-mRNA 3’-end processing. It is demonstrated that the interaction of ATP and Pcf11 with Clp1 are linked events: Loss of Clp1-ATP binding results in the abrogation of the Pcf11-Clp1 interaction and leads to Clp1 instability in vitro, and similarly, mutations that directly uncouple the Pcf11-Clp1 interaction also disrupt Clp1-ATP binding and cause Clp1 instability in vitro. An in vivo mutational analysis in S. cerevisiae revealed that both Clp1-ATP binding and the Pcf11-Clp1 interaction are essential for yeast survival. Further cell and immunoprecipitation studies demonstrated that one essential function of Clp1 is as a chaperone of Pcf11, and RT-qPCR analysis of mRNA from a sample set of yeast genes points to a role for these proteins in transcription and transcription termination rather than in poly(A) site selection.572University College London (University of London)http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.568289http://discovery.ucl.ac.uk/1369193/Electronic Thesis or Dissertation |
collection |
NDLTD |
sources |
NDLTD |
topic |
572 |
spellingShingle |
572 Hedden, J. J. The role of Clp1 and Pcf11 in transcription and pre-mRNA 3’-end processing |
description |
Eukaryotic transcripts require a number of complex cotranscriptional modifications and processing events before translation to protein. Clp1 and Pcf11 are subunits of cleavage factor IA (CFIA), an essential component of the Saccharomyces cerevisiae pre-mRNA 3’-end processing machinery. The crystal structure of a Clp1-Pcf11 complex was determined previously and revealed the binding of ATP to a highly-conserved P-loop motif and a tight Pcf11-Clp1 interaction facilitated by a number of highly-conserved Pcf11 residues. Nonetheless, the biological function of both Clp1-ATP binding and the Pcf11-Clp1 interaction was not well understood. The work in this thesis combines an in vitro and in vivo investigation of the Clp-ATP and Clp-Pcf11 interactions in an effort to understand the function of these factors in transcription and pre-mRNA 3’-end processing. It is demonstrated that the interaction of ATP and Pcf11 with Clp1 are linked events: Loss of Clp1-ATP binding results in the abrogation of the Pcf11-Clp1 interaction and leads to Clp1 instability in vitro, and similarly, mutations that directly uncouple the Pcf11-Clp1 interaction also disrupt Clp1-ATP binding and cause Clp1 instability in vitro. An in vivo mutational analysis in S. cerevisiae revealed that both Clp1-ATP binding and the Pcf11-Clp1 interaction are essential for yeast survival. Further cell and immunoprecipitation studies demonstrated that one essential function of Clp1 is as a chaperone of Pcf11, and RT-qPCR analysis of mRNA from a sample set of yeast genes points to a role for these proteins in transcription and transcription termination rather than in poly(A) site selection. |
author |
Hedden, J. J. |
author_facet |
Hedden, J. J. |
author_sort |
Hedden, J. J. |
title |
The role of Clp1 and Pcf11 in transcription and pre-mRNA 3’-end processing |
title_short |
The role of Clp1 and Pcf11 in transcription and pre-mRNA 3’-end processing |
title_full |
The role of Clp1 and Pcf11 in transcription and pre-mRNA 3’-end processing |
title_fullStr |
The role of Clp1 and Pcf11 in transcription and pre-mRNA 3’-end processing |
title_full_unstemmed |
The role of Clp1 and Pcf11 in transcription and pre-mRNA 3’-end processing |
title_sort |
role of clp1 and pcf11 in transcription and pre-mrna 3’-end processing |
publisher |
University College London (University of London) |
publishDate |
2012 |
url |
http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.568289 |
work_keys_str_mv |
AT heddenjj theroleofclp1andpcf11intranscriptionandpremrna3endprocessing AT heddenjj roleofclp1andpcf11intranscriptionandpremrna3endprocessing |
_version_ |
1718393286205374464 |