title: The human hyaluronan synthase 2 (HAS2) gene and its natural antisense RNA exhibit coordinated expression in the renal proximal tubular epithelial cell.
abstract: Aberrant expression of the human hyaluronan synthase 2 (HAS2) gene has been implicated in the pathology of malignancy, pulmonary arterial hypertension, osteoarthritis, asthma, thyroid dysfunction, and large organ fibrosis. Renal fibrosis is associated with increased cortical synthesis of hyaluronan (HA), an extracellular matrix glycosaminoglycan, and we have shown that HA is a correlate of interstitial fibrosis in vivo. Our previous in vitro data have suggested that both HAS2 transcriptional induction and subsequent HAS2-driven HA synthesis may contribute to kidney fibrosis via phenotypic modulation of the renal proximal tubular epithelial cell (PTC). Post-transcriptional regulation of HAS2 mRNA synthesis by the natural antisense RNA HAS2-AS1 has recently been described in osteosarcoma cells, but the antisense transcript was not detected in kidney. In this study, PTC stimulation with IL-1β or TGF-β1 induced coordinated temporal profiles of HAS2-AS1 and HAS2 transcription. Constitutive activity of the putative HAS2-AS1 promoter was demonstrated, and transcription factor-binding sequence motifs were identified. Knockdown of Sp1/Sp3 expression by siRNA blunted IL-1β induction of both HAS2-AS1 and HAS2, and Smad2/Smad3 knockdown similarly attenuated TGF-β1 stimulation. Inhibition of IL-1β-stimulated HAS2-AS1 RNA induction using HAS2-AS1-specific siRNAs also suppressed up-regulation of HAS2 mRNA transcription. The thermodynamic feasibility of HAS2-AS1/HAS2 heterodimer formation was demonstrated in silico, and locus-specific cytoplasmic double-stranded RNA was detected in vitro. In summary, our data show that transcriptional induction of HAS2-AS1 and HAS2 occurs simultaneously in PTCs and suggest that transcription of the antisense RNA stabilizes or augments HAS2 mRNA expression in these cells via RNA/mRNA heteroduplex formation.
authors.affiliation: Institute of Nephrology, Cardiff University School of Medicine, Heath Park, Cardiff, UK.
authors.forename: Daryn R
authors.initials: DR
authors.lastname: Michael
authors.forename: Aled O
authors.initials: AO
authors.lastname: Phillips
authors.forename: Aleksandra
authors.initials: A
authors.lastname: Krupa
authors.forename: John
authors.initials: J
authors.lastname: Martin
authors.forename: James E
authors.initials: JE
authors.lastname: Redman
authors.forename: Abdalsamed
authors.initials: A
authors.lastname: Altaher
authors.forename: Rachel D
authors.initials: RD
authors.lastname: Neville
authors.forename: Jason
authors.initials: J
authors.lastname: Webber
authors.forename: Min-young
authors.initials: MY
authors.lastname: Kim
authors.forename: Timothy
authors.initials: T
authors.lastname: Bowen
mesh_terms: D045744:Cell Line, Tumor; D004847:Epithelial Cells / Q000378:metabolism* / Q000473:pathology; D005355:Fibrosis; D005786:Gene Expression Regulation; D055785:Gene Knockdown Techniques; D014453:Glucuronosyltransferase / Q000096:biosynthesis* / Q000235:genetics; D006801:Humans; D000076002:Hyaluronan Synthases; D006820:Hyaluronic Acid / Q000096:biosynthesis; D006976:Hypertension, Pulmonary / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D053583:Interleukin-1beta / Q000235:genetics / Q000378:metabolism; D007674:Kidney Diseases / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D007687:Kidney Tubules, Proximal / Q000378:metabolism* / Q000473:pathology; D009692:Nucleic Acid Heteroduplexes / Q000096:biosynthesis / Q000235:genetics; D016372:RNA, Antisense / Q000096:biosynthesis* / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D020218:Response Elements / Q000235:genetics; D051899:Smad2 Protein / Q000235:genetics / Q000378:metabolism; D051900:Smad3 Protein / Q000235:genetics / Q000378:metabolism; D016329:Sp1 Transcription Factor / Q000235:genetics / Q000378:metabolism; D051705:Sp3 Transcription Factor / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic; D053773:Transforming Growth Factor beta1 / Q000235:genetics / Q000378:metabolism
chemical_list: D053583:Interleukin-1beta; D009692:Nucleic Acid Heteroduplexes; D016372:RNA, Antisense; D012333:RNA, Messenger; C497782:SMAD2 protein, human; C497790:SMAD3 protein, human; C497121:SP3 protein, human; D051899:Smad2 Protein; D051900:Smad3 Protein; D016329:Sp1 Transcription Factor; D053773:Transforming Growth Factor beta1; D051705:Sp3 Transcription Factor; D006820:Hyaluronic Acid; D014453:Glucuronosyltransferase; C101754:HAS2 protein, human; D000076002:Hyaluronan Synthases