Protein synthesis and urokinase mRNA metabolism.

Expression of the urokinase-type plasminogen activator (uPA) is under tight regulation by hormones, cytokines and growth factors under physiological conditions. Treatment of lung epithelial (Beas2B) cells with translation inhibitors induces uPA mRNA expression, as well as early response genes. To understand the specific expression and regulation of uPA mRNA, we treated Beas2B cells with cycloheximide (CycD), anisomycin, emitine and puromycin in a time-dependent manner and measured uPA mRNA expression by Northern blotting. All these agents induced uPA mRNA by two- to seven-fold within 3 h after treatment in Beas2B cells. CycD, emitine, puromycin and anisomycin also enhanced uPA mRNA half-life by three- to five-fold in Beas2B cells treated with DRB, an inhibitor of transcription. However, run-on-transcription experiments indicated that these agents failed to induce uPA mRNA transcription indicating that they augment uPA mRNA mainly due to increased stability. Using gel mobility shift, we identified an uPA mRNA binding protein (uPA mRNABp) that selectively binds to uPA mRNA [Gyetko MR, Todd III RF, Wilkinson CC, Sitrin RG: The urokinase receptor is required for human monocyte chemotaxis in vitro. J Clin Invest 93: 1380-1387, 1994]. Binding of both cytoplasmic and nuclear uPA mRNABp to uPA mRNA was abolished after treatment with translation inhibitors, which coincides with the maximal expression of uPA mRNA. We also found a similar decline in HuR and heterogeneous nuclear ribonucleoprotein C (hnRNPC) which are known to stabilize uPA mRNA both in the nuclear and cytosolic compartments. These results strongly suggest that increased uPA mRNA stability induced by translational inhibitors involves the interaction of uPA mRNA with a degrading protein factor rather than increased interaction of proteins that are known to stabilize uPA mRNA. These data also strongly suggests that down-regulation of the uPA-uPA mRNABp interaction by translational inhibitors rather than the translocation of uPA mRNABp contributes to increased uPA mRNA stability. This pathway may regulate uPA-mediated functions of the lung epithelium in the context of inflammation or neoplasia.