A region within the 5′-untranslated region of hypoxia-inducible factor-1α mRNA mediates its turnover in lung adenocarcinoma cells

Abstract

Previously, we showed that CL1-5 cells express more hypoxia-inducible factor-1α (HIF-1α) than the parental CL1 cells, which bestows CL1-5 cells a stronger invasive activity. Here, we investigated the mechanisms underlying the differential expression of HIF-1α mRNA in CL1 and CL1-5 cells. Data showed that the transcription rate of HIF-1α gene in CL1 cells was slightly higher than that of CL1-5 cells, suggesting that the expression of HIF-1α mRNA in CL1 cells was repressed by post-transcriptional mechanisms. RNA electrophoretic mobility shift assays revealed a 61-base segment (designated as D5) within the 5′-untranslated repeat of HIF-1α mRNA, with which the CL1 cell lysates formed more prominent complexes (including complex I) than did CL1-5 cell lysates. Insertion of D5 into a reporter construct reduced the half-life of the chimeric transcripts in transfected CL1 but not CL1-5 cells; conversely, overexpression of D5-containing reporter construct in CL1 cells increased HIF-1α mRNA. We also identified the polypyrimidine tract-binding protein (PTB) as a required component of complex I. Deletion of the RNA recognition motif 1 (RRM1) or RRM3 of PTB abolished the formation of complex I. Our data showed that CL1 cells expressed more PTB than CL1-5 cells. Inhibition of PTB expression in CL1 cells decreased the formation of complex I, whereas overexpression of PTB in CL1-5 cells increased the levels of complex I, decreased the stability of HIF-1α and D5-containing chimeric mRNAs, and decreased cell invasiveness. In sum, we have identified in lung adenocarcinoma cells a mechanism that regulates HIF-1α expression by modulating HIF-1α mRNA stability. © 2009 by The American Society for Biochemistry and Molecular Biology, Inc.