Posttranscriptional regulation of urokinase receptor mRNA: identification of a novel urokinase receptor mRNA binding protein in human mesothelioma cells.
Treatment of human pleural mesothelioma (MS-1) cells with phorbol myristate acetate (PMA) and cycloheximide results in 17- and 10-fold, respectively, increases in steady-state expression of urokinase-type plasminogen activator receptor (uPAR) mRNA. Studies of transcriptional inhibition by actinomycin D showed four- and sixfold extensions of uPAR mRNA half-life in MS-1 cells treated with PMA and cycloheximide, respectively, suggesting that uPAR gene expression involves a posttranscriptional regulatory mechanism. Using gel mobility shift and UV cross-linking assays, we identified a 50-kDa uPAR mRNA binding protein (uPAR mRNABp) that selectively bound to a 51-nucleotide (nt) fragment of mRNA corresponding to the uPAR coding region. We investigated the possibility that this 51-nt protein binding fragment of uPAR mRNA contains regulatory information for message stability. Chimeric beta-globin/uPAR/beta-globin mRNA containing the 51-nt protein binding fragment was able to destabilize otherwise stable beta-globin mRNA. Conversely, a control chimeric beta-globin/uPAR/beta-globin mRNA containing a 51-nt fragment of the uPAR coding region that does not bind uPAR mRNABp was stable under identical conditions. Binding of uPAR mRNABp to uPAR mRNA was abolished after treatment with cycloheximide and rapidly down-regulated by PMA. These data suggest that the 51-nt protein binding fragment of uPAR mRNA may be involved in mRNA turnover as well as in cycloheximide-induced uPAR message stabilization. Our results indicate a novel mechanism of uPAR gene regulation in which cis elements within a 51-nt coding region interact with a uPAR mRNABp to regulate uPAR message stability.
Shetty
S
S
Department of Medicine, University of Texas Health Science Center, Tyler 75710, USA.
Kumar
A
A
Idell
S
S
D001483:Base Sequence; D001665:Binding Sites; D003513:Cycloheximide / Q000494:pharmacology; D003609:Dactinomycin / Q000494:pharmacology; D004847:Epithelial Cells; D004848:Epithelium / Q000378:metabolism; D015870:Gene Expression; D005914:Globins / Q000235:genetics; D006801:Humans; D008070:Lipopolysaccharides / Q000494:pharmacology; D008654:Mesothelioma / Q000378:metabolism*; D008969:Molecular Sequence Data; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D010997:Pleural Neoplasms / Q000378:metabolism*; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D012313:RNA / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000235:genetics*; D055293:Receptors, Urokinase Plasminogen Activator; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D016212:Transforming Growth Factor beta / Q000494:pharmacology; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D014568:Urokinase-Type Plasminogen Activator
D008070:Lipopolysaccharides; D019384:Nucleic Acid Synthesis Inhibitors; C524898:PLAUR protein, human; D011500:Protein Synthesis Inhibitors; D012333:RNA, Messenger; C039919:RNA, recombinant; D016601:RNA-Binding Proteins; D011956:Receptors, Cell Surface; D055293:Receptors, Urokinase Plasminogen Activator; D016212:Transforming Growth Factor beta; D014409:Tumor Necrosis Factor-alpha; D003609:Dactinomycin; D012313:RNA; D005914:Globins; D003513:Cycloheximide; D014568:Urokinase-Type Plasminogen Activator; D013755:Tetradecanoylphorbol Acetate