Novel MicroRNA Reporter Uncovers Repression of Let-7 by GSK-3β

Several members of the let-7 microRNA family are downregulated in ovarian and other cancers. They are thought to act as tumor suppressors by lowering growth-promoting and anti-apoptotic proteins. In order to measure cellular let-7 levels systematically, we have developed a highly sensitive let-7 rep...

Full description

Bibliographic Details
Main Authors:	Guo, Rong, Abdelmohsen, Kotb, Morin, Patrice J., Gorospe, Myriam
Format:	Online
Language:	English
Published:	Public Library of Science 2013
Online Access:	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3694080/

id	pubmed-3694080
recordtype	oai_dc
spelling	pubmed-36940802013-07-09 Novel MicroRNA Reporter Uncovers Repression of Let-7 by GSK-3β Guo, Rong Abdelmohsen, Kotb Morin, Patrice J. Gorospe, Myriam Research Article Several members of the let-7 microRNA family are downregulated in ovarian and other cancers. They are thought to act as tumor suppressors by lowering growth-promoting and anti-apoptotic proteins. In order to measure cellular let-7 levels systematically, we have developed a highly sensitive let-7 reporter assay system based on the expression of a chimeric mRNA that contains the luciferase coding region and a 3′-untranslated region (UTR) bearing two let-7-binding sites. In cells expressing the reporter construct, termed pmirGLO-let7, luciferase activity was high when let-7 was absent, while luciferase activity was low when let-7 levels were elevated. The ovarian cancer cell lines BG-1 and UCI-101 were transfected with the let-7 reporter and surveyed with a library of kinase inhibitors in order to identify pathways affecting let-7 activity. Among the inhibitors causing changes in endogenous let-7 abundance, the lowering of glycogen synthase kinase 3 (GSK-3)β function specifically increased let-7 levels and lowered luciferase activity. Similarly, silencing GSK-3β increased both mature and primary-let-7 levels in BG-1 cells, and decreased BG-1 cell survival. Further studies identified p53 as a downstream effector of the GSK-3β-mediated repression of let-7 biosynthesis. Our studies highlight GSK-3β as a novel therapeutic target in ovarian tumorigenesis. Public Library of Science 2013-06-26 /pmc/articles/PMC3694080/ /pubmed/23840442 http://dx.doi.org/10.1371/journal.pone.0066330 Text en https://creativecommons.org/publicdomain/zero/1.0/ This is an open-access article distributed under the terms of the Creative Commons Public Domain declaration, which stipulates that, once placed in the public domain, this work may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose.
repository_type	Open Access Journal
institution_category	Foreign Institution
institution	US National Center for Biotechnology Information
building	NCBI PubMed
collection	Online Access
language	English
format	Online
author	Guo, Rong Abdelmohsen, Kotb Morin, Patrice J. Gorospe, Myriam
spellingShingle	Guo, Rong Abdelmohsen, Kotb Morin, Patrice J. Gorospe, Myriam Novel MicroRNA Reporter Uncovers Repression of Let-7 by GSK-3β
author_facet	Guo, Rong Abdelmohsen, Kotb Morin, Patrice J. Gorospe, Myriam
author_sort	Guo, Rong
title	Novel MicroRNA Reporter Uncovers Repression of Let-7 by GSK-3β
title_short	Novel MicroRNA Reporter Uncovers Repression of Let-7 by GSK-3β
title_full	Novel MicroRNA Reporter Uncovers Repression of Let-7 by GSK-3β
title_fullStr	Novel MicroRNA Reporter Uncovers Repression of Let-7 by GSK-3β
title_full_unstemmed	Novel MicroRNA Reporter Uncovers Repression of Let-7 by GSK-3β
title_sort	novel microrna reporter uncovers repression of let-7 by gsk-3β
description	Several members of the let-7 microRNA family are downregulated in ovarian and other cancers. They are thought to act as tumor suppressors by lowering growth-promoting and anti-apoptotic proteins. In order to measure cellular let-7 levels systematically, we have developed a highly sensitive let-7 reporter assay system based on the expression of a chimeric mRNA that contains the luciferase coding region and a 3′-untranslated region (UTR) bearing two let-7-binding sites. In cells expressing the reporter construct, termed pmirGLO-let7, luciferase activity was high when let-7 was absent, while luciferase activity was low when let-7 levels were elevated. The ovarian cancer cell lines BG-1 and UCI-101 were transfected with the let-7 reporter and surveyed with a library of kinase inhibitors in order to identify pathways affecting let-7 activity. Among the inhibitors causing changes in endogenous let-7 abundance, the lowering of glycogen synthase kinase 3 (GSK-3)β function specifically increased let-7 levels and lowered luciferase activity. Similarly, silencing GSK-3β increased both mature and primary-let-7 levels in BG-1 cells, and decreased BG-1 cell survival. Further studies identified p53 as a downstream effector of the GSK-3β-mediated repression of let-7 biosynthesis. Our studies highlight GSK-3β as a novel therapeutic target in ovarian tumorigenesis.
publisher	Public Library of Science
publishDate	2013
url	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3694080/
_version_	1611989868398772224

Novel MicroRNA Reporter Uncovers Repression of Let-7 by GSK-3β

Similar Items