, Volume 272, Issue 1-2, pp 107-118

Regulation of urokinase receptor mRNA stability by hnRNP C in lung epithelial cells

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Abstract

Increased urokinase receptor (uPAR) expression as well as stabilisation of uPAR mRNA contribute to the pathogenesis of lung inflammation and neoplasia. Post-transcriptional regulation of uPAR mRNA involves interaction of both coding and 3′-UTR sequences with regulatory uPAR mRNA binding proteins (Bps). In order to identify novel regulatory interactions, we performed gel mobility shift and UV cross-linking assays and found two distinct uPAR mRNA-protein complexes. We identified a rapidly migrating 40 kDa uPAR mRNABp that selectively bound a 110 nucleotide (nt) fragment of the uPAR mRNA 3′UTR. Chimeric β-globin/uPAR mRNA containing the 110 nt 40 kDa protein binding fragment destabilised stable β-globin mRNA with a rate of decay identical to that of chimeric β-globin/uPAR containing the full uPAR 3′UTR. The 40 kDa uPAR 3′UTR Bp was purified using poly (U) sepharose and identified as heterogeneous nuclear ribonucleoprotein C (hnRNPC). Finally, we confirmed its interaction with the uPAR mRNA 3′ UTR by gel mobility supershift assay using an anti-hnRNPC antibody. Direct in vivo interaction of hnRNPC with the uPAR mRNA 3′UTR was demonstrated by immunoprecipitation and combined RT PCR-Southern blotting assay. Co-transfection of hnRNPC cDNA in Beas2B cells reversed destabilisation of chimeric β-globin/uPAR 3′UTR mRNA and its over-expression also induced uPAR protein and mRNA expression through stabilisation of uPAR mRNA. These observations indicate a novel mechanism of uPAR gene regulation in lung epithelial cells in which cis elements within a 110 nt uPAR mRNA 3′UTR sequence interact with hnRNPC to regulate uPAR mRNA stability.