Regulation of fibroblast growth factor-2 by an endogenous antisense RNA and by argonaute-2.
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.
MacFarlane
Leigh-Ann
LA
Department of Physiology and Biophysics, Faculty of Medicine, Dalhousie University, Halifax, Nova Scotia, Canada.
Gu
Ying
Y
Casson
Alan G
AG
Murphy
Paul R
PR
D020413:3' Untranslated Regions / Q000235:genetics; D060565:Argonaute Proteins; D015153:Blotting, Western; D045744:Cell Line, Tumor; D015852:Eukaryotic Initiation Factor-2 / Q000235:genetics / Q000502:physiology*; D016222:Fibroblast Growth Factor 2 / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D008957:Models, Genetic; D020411:Oligonucleotide Array Sequence Analysis; D016372:RNA, Antisense / Q000235:genetics / Q000502:physiology*; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics / Q000502:physiology; D020133:Reverse Transcriptase Polymerase Chain Reaction
D020413:3' Untranslated Regions; C466422:AGO2 protein, human; D060565:Argonaute Proteins; D015852:Eukaryotic Initiation Factor-2; D016372:RNA, Antisense; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D016222:Fibroblast Growth Factor 2