Open AccessRegulation of Fibroblast Growth Factor-2 by an Endogenous Antisense RNA and by Argonaute-2
Author(s) -
Leigh-Ann MacFarlane,
Ying Gu,
Alan G. Casson,
Paul R. Murphy
Publication year - 2010
Publication title -
molecular endocrinology
Language(s) - English
Resource type - Journals
eISSN - 1944-9917
pISSN - 0888-8809
DOI - 10.1210/me.2009-0367
Subject(s) - gene knockdown , biology , small interfering rna , microbiology and biotechnology , argonaute , antisense rna , rna interference , rna , messenger rna , untranslated region , rna binding protein , sense (electronics) , trans acting sirna , cell culture , gene , genetics , chemistry
We have previously reported that elevated fibroblast growth factor-2 (FGF-2) expression is associated with tumor recurrence and reduced survival after surgical resection of esophageal cancer and that these risks are reduced in tumors coexpressing an endogenous antisense (FGF-AS) RNA. In the present study, we examined the role of the endogenous FGF-AS transcript in the regulation of FGF-2 expression in the human lung adenocarcinoma cell line Seg-1. FGF-2 and FGF-AS were temporally and spatially colocalized in the cytoplasm of individual cells, and knockdown of either FGF-2 or FGF-AS by target-specific siRNAs resulted in dose-dependent up-regulation of the complementary transcript and its encoded protein product. Using a luciferase reporter system, we show that these effects are mediated by interaction of the endogenous antisense RNA with the 3'-untranslated region of the FGF-2 mRNA. Deletion mapping identified a 392-nucleotide sequence in the 5823-nucleotide FGF-2 untranslated tail that is targeted by FGF-AS. Small interfering RNA-mediated knockdown of either FGF-AS or FGF-2 significantly increased the stability of the complementary partner mRNA, demonstrating that these mRNAs are mutually regulatory. Knockdown of FGF-AS also resulted in reduced expression of argonaute-2 (AGO-2) and a number of other elements of the endogenous micro-RNA/RNA interference pathways. Conversely, small interfering RNA-mediated knockdown of AGO-2 significantly increased the stability of the FGF-2 mRNA transcript and the steady-state levels of both FGF-2 mRNA and protein, suggesting a role for AGO-2 in the regulation of FGF-2 expression.