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Tailing-dependent trimming of RISC-associated miRNAs by th HuR protein

Sokol, Lena. Tailing-dependent trimming of RISC-associated miRNAs by th HuR protein. 2014, PhD Thesis, University of Basel, Faculty of Science.

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Official URL: http://edoc.unibas.ch/diss/DissB_10965

Abstract

HuR is a ubiquitously expressed AU-rich element (ARE) binding protein. AREs are regulatory, typically destabilizing sequences found in the 3' UTR of many mRNAs in eukaryotes. However, binding of HuR acts to stabilize these messages. Another type of regulatory elements, miRNA-binding sequences are also found on the mRNA 3'UTR. miRNAs are short, (~22 nt) non-coding RNAs which guide the RISC complex to regulate the expression of proteins involved in the regulation of numerous biological processes.
HuR and miRNA sites have been shown to often co-localize on target mRNAs, and several studies have shown that there is a cross-talk between HuR and miRNAs. For example, HuR can counteract miR-122-mediated repression of CAT-1 mRNA, a process which is accompanied by Ago displacement from mRNA.
It was previously shown that HuR possesses an RNA 3'-terminal adenosyl transferase activity; however, the physiological substrates were not determined. In this thesis, we report that miRNAs are bound by HuR and act as substrates for the HuR-mediated transferase activity and that HuR can polyadenylate miRNAs. We further describe different type of evidence strongly suggesting that HuR also has a 3'-5' exonuclease activity acting on miRNAs, and describe and test in vitro a model of how HuR antagonizes the Ago-bound miRNA associated with mRNA. To access the miRNA-Ago complex associated in cis with the same target RNA, HuR recognizes the miRNA 3' end and adds to it a poly(A) tail, thus potentially weakening the Ago-miRNA interaction and creating a landing pad for the nuclease, which in turn leads to exonucleolytic degradation of miRNA turnover and displacement of Ago from RNA.
We further show that HuR knockdown in HCT116 colon carcinoma cells has a differential effect on mature miRNAs as compared to miRNA isoforms containing 3'-terminal non-templated additions of A residues. The levels of these isoforms are reduced upon HuR depletion, suggesting that HuR promotes miRNA tailing and degradation also in vivo.
Advisors:Filipowicz, Witold
Committee Members:Martinez, Javier
Faculties and Departments:09 Associated Institutions > Friedrich Miescher Institut FMI > Biochemistry and function of RNA interference and microRNAs (Filipowicz)
Item Type:Thesis
Thesis no:10965
Bibsysno:Link to catalogue
Number of Pages:142 Bl.
Language:English
Identification Number:
Last Modified:30 Jun 2016 10:56
Deposited On:23 Dec 2014 14:30

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