Year: 2014
Type: dissertation
Abstract: Vts1p is a member of the Smaug family of sequence-specific RNA binding proteins that interact with stem-loop sequences to degrade and/or translationally repress target transcripts. To identify the molecular mechanism by which Vts1p represses target transcripts, I have investigated the role of one of Vts1p’s conserved domains, the SSR1 domain, in Vts1p-mediated repression. In vitro SSR1 functions as a dimerization domain and I provide evidence that SSR1 mediates dimerization in vivo. I have created Vts1p mutants which abolish dimerization and found that dimerization is required for full Vts1p-mediated repression and robust destabilization of a reporter containing two SREs. My data supports a model whereby dimerization of Vts1p functions to enhance binding of Vts1p to a target transcript carrying two or more SREs. Analysis of putative Vts1p mRNA targets suggests that dimerization may be important for proper repression of many target transcripts since a majority contain multiple SREs. I have also shown that Vts1p interacts with the Ccr4p/Pop2p/Not deadenylase in an RNA independent fashion. This is consistent with the model that Vts1p recruits the Ccr4p/Pop2p/Not deadenylase to target transcripts to induce deadenylation and subsequent degradation. Finally, by tethering Vts1p to a reporter construct, I have shown that two separate regions of Vts1p, the N-terminus and the C-terminus, are sufficient for repression, suggesting that Vts1p employs multiple mechanisms to regulate the expression of target mRNAs. I have also shown that the N-terminus contains a conserved motif that functions in Vts1p-mediated translational repression while playing no role in transcript decay. This is the first evidence that Vts1p mediates the translational repression of a target transcript.
Source: TSpace (University of Toronto)
Author: Melissa A. Bieman
Language: English
Topic: RNA Research and Splicing
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W3155214981