Regulation of human inducible nitric oxide synthase expression by an upstream open reading frame

Publication date: Available online 18 April 2019Source: Nitric OxideAuthor(s): Fabian Gather, Katja Schmitz, Kathrin Koch, Lea-Marie Vogt, Andrea Pautz, Hartmut KleinertAbstractThe human inducible nitric oxide synthase (iNOS) gene contains an upstream open reading frame (uORF) in its 5′-untranslated region (5′-UTR) implying a translational regulation of iNOS expression. Transfection experiments in human DLD-1 cells revealed that the uORF although translatable seems not to inhibit the translation start at the bona fide ATG. Our data clearly show that human iNOS translation is cap-dependent and that the 5′-UTR of the iNOS mRNA contains no internal ribosome entry site. Translation of the bona fide coding sequence is most likely mediated by a leaky scanning mechanism.The 5′-UTR is encoded by exon 1 and exon 2 of the iNOS gene with the uORF stop codon located in front of the first intron indicating an involvement of the nonsense mediated RNA decay (NMD) in iNOS regulation. SiRNA-mediated down-regulation of Upf1 resulted in enhanced endogenous cytokine iNOS expression in human DLD-1 cells. Transfection of constructs containing iNOS exon 1, intron 1 and exon 2 in front of a luciferase gene showed a clear effect of the mutation of the uORF-ATG on luciferase reportergene expression.Our data indicate that the uORF in the 5′-UTR sequence of human iNOS gene reduces its expression via the NMD mechanism.

Source: Nitric Oxide - April 19, 2019 Category: Chemistry Source Type: research