Complex contribution of the 3 '-untranslated region to the expressional regulation of the human inducible nitric-oxide synthase gene - Involvement ofthe RNA-binding protein HuR

Cytokine stimulation of human DLD-1 cells resulted in a marked expression o f nitric-oxide synthase (NOS) II mRNA and protein accompanied by only a mod erate increase in transcriptional activity. Also, there was a basal transcr iption of the NOS II gene, which did not result in measurable NOS II expres sion. The 3'-untranslated region (3'-UTR) of the NOS II mRNA contains four AUUUA motifs and one AUUUUA motif, known to destabilize the mRNAs of proto- oncogenes, nuclear transcription factors, and cytokines. Luciferase reporte r gene constructs containing the NOS II 3'-UTR showed a significantly reduc ed luciferase activity. The embryonic lethal abnormal vision (ELAV)-like pr otein HuR was found to bind with high affinity to the adenylate/uridylate-r ich elements of the NOS II 3'-UTR. Inhibition of HuR with antisense constru cts reduced the cytokine-induced NOS II mRNA, whereas overexpression of HuR potentiated the cytokine-induced NOS II expression. This provides evidence that NOS II expression is regulated at the transcriptional and post-transc riptional level. Binding of HuR to the 3'-UTR of the NOS II mRNA seems to p lay an essential role in the stabilization of this mRNA.