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RNAi-mediated knockdown of the Rhau helicase preferentially depletes proteins with a Guanine-quadruplex motif in the 5'-UTR of their mRNA

Author:
Vester, Karen, Eravci, Murat, Serikawa, Tatsuo, Schütze, Tatjana, Weise, Christoph, Kurreck, Jens
Source:
Biochemical and biophysical research communications 2019 v.508 no.3 pp. 756-761
ISSN:
0006-291X
Subject:
5' untranslated regions, RNA helicases, bioinformatics, data collection, gene expression, gene expression regulation, guanine, humans, liquid chromatography, nucleic acid conformation, protein content, proteins, proto-oncogenes, tandem mass spectrometry
Abstract:
Guanine-quadruplex (G-quadruplex) structures in mRNAs have been shown to modulate gene expression. However, the overall biological relevance of this process is under debate, as cellular helicases unwind G-quadruplex structures. The helicase Rhau (encoded by the DHX36 gene) was reported to be the major source of RNA G-quadruplex resolving activity in lysates of human cells. In the current study, we depleted Rhau by RNAi-mediated silencing and analyzed the effect on proteins whose mRNAs harbor a G-quadruplex motif in their 5'-UTRs. A targeted investigation of the proto-oncogenes Bcl-2 and NRAS, which are well-known examples for the translational repression of G-quadruplex structures, did not reveal effects caused by Rhau silencing. We therefore carried out a global analysis of changes in protein levels by label-free quantification using liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS). Following Rhau knockdown, of all the identified proteins, only 1.9% were significantly downregulated to at least 70%. According to a bioinformatic analysis with the QGRS mapper, 33% of the downregulated proteins were predicted to harbor a G-quadruplex motif in the 5'-UTR of their respective mRNAs, compared to only 11% in the complete dataset. This indicates that in an unexpectedly small set of genes, in which G-quadruplex motifs are unusually common in the 5'-UTR of their mRNAs, Rhau helicase is responsible for the regulation of their expression.
Agid:
6250510