title: Biogenesis of short intronic repeat 27-nucleotide small RNA from endothelial nitric-oxide synthase gene.
abstract: Endothelial nitric-oxide synthase (eNOS) is a constitutively expressed gene in endothelium that produces NO and is critical for vascular integrity. Previously, we reported that the 27-nucleotide (nt) repeat polymorphism in eNOS intron 4, a source of 27-nt small RNA, which inhibits eNOS expression, were associated with cardiovascular risk and expression of the eNOS gene. In the current study, we investigated the biogenesis of the intron 4-derived 27-nt small RNA. Using Northern blot, we showed that the eNOS-derived 27-nt short intronic repeat RNA (sir-RNA) expressed only in the eNOS expressing endothelial cells. Cells containing 10 x 27- or 5 x 27-nt repeats produced higher levels of 27nt sir-RNA and lower levels of eNOS mRNA than the cells with 4 x 27-nt repeats. The 27nt sir-RNA was mostly present within the endothelial nuclei. When the splicing junctions of the 27-nt repeat containing intron 4 in the full-length eNOS cDNA vector were mutated, 27nt sir-RNA biogenesis was abolished. Suppression of Drosha or Dicer diminished the biogenesis of the 27nt sir-RNA. Our study suggests that the 27nt sir-RNA derived through eNOS pre-mRNA splicing may represent a new class of small RNA. The more eNOS is transcribed or higher number of the 27-nt repeats, the more 27nt sir-RNA is produced, which functions as a negative feedback self-regulator by specifically inhibiting the host gene eNOS expression. This novel molecular model may be responsible for quantitative differences between individuals carrying different numbers of the polymorphic repeats hence the cardiovascular risk.
authors.affiliation: Adult Section of Cardiothoracic Surgery, Texas Heart Institute at St Luke's Episcopal Hospital, Division of Cardiothoracic Surgery, Michael E DeBakey Department of Surgery, Baylor College of Medicine, Houston, TX 77030, USA. mzhang1@bcm.edu
authors.forename: Ming-Xiang
authors.initials: MX
authors.lastname: Zhang
authors.forename: Cheng
authors.initials: C
authors.lastname: Zhang
authors.forename: Ying H
authors.initials: YH
authors.lastname: Shen
authors.forename: Jian
authors.initials: J
authors.lastname: Wang
authors.forename: Xiao Nan
authors.initials: XN
authors.lastname: Li
authors.forename: Yun
authors.initials: Y
authors.lastname: Zhang
authors.forename: Joseph
authors.initials: J
authors.lastname: Coselli
authors.forename: Xing Li
authors.initials: XL
authors.lastname: Wang
mesh_terms: D017398:Alternative Splicing / Q000235:genetics; D001483:Base Sequence; D002478:Cells, Cultured; D018076:DNA, Complementary / Q000235:genetics; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D007438:Introns / Q000235:genetics*; D008969:Molecular Sequence Data; D052250:Nitric Oxide Synthase Type III / Q000235:genetics* / Q000378:metabolism*; D009711:Nucleotides / Q000096:biosynthesis* / Q000235:genetics*; D009928:Organ Specificity; D011401:Promoter Regions, Genetic / Q000235:genetics; D012313:RNA / Q000096:biosynthesis* / Q000235:genetics*
chemical_list: D018076:DNA, Complementary; D009711:Nucleotides; D012313:RNA; D052250:Nitric Oxide Synthase Type III