The Human Hyaluronan Synthase 2 (HAS2) Gene and Its Natural Antisense RNA Exhibit Coordinated Expression in the Renal Proximal Tubular Epithelial Cell
The Human Hyaluronan Synthase 2 (HAS2) Gene and Its Natural Antisense RNA Exhibit Coordinated Expression in the Renal Proximal Tubular Epithelial Cell
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DOI：
10.1074/jbc.m111.233916
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发表时间：
2011-06-03
影响因子：
4.8
通讯作者：
Bowen, Timothy
中科院分区：
文献类型：
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作者：
Michael, Daryn R.;Phillips, Aled O.;Bowen, Timothy
人类透明质酸合成酶2 （HAS2）基因的异常表达与恶性肿瘤、肺动脉高压、骨关节炎、哮喘、甲状腺功能障碍和大器官纤维化的病理有关。肾纤维化与皮质透明质酸（HA）的合成增加有关，HA是一种细胞外基质糖胺聚糖，我们已经证明HA与体内间质纤维化有关。我们之前的体外数据表明，HAS2转录诱导和随后的HAS2驱动的HA合成可能通过肾近端小管上皮细胞（PTC）的表型调节促进肾纤维化。天然反义RNA HAS2- as1对HAS2 mRNA合成的转录后调控最近在骨肉瘤细胞中被描述，但在肾脏中未检测到反义转录物。在这项研究中，用IL-1 β或tgf - β 1刺激PTC可诱导HAS2- as1和HAS2转录的协调时间谱。证明了推定的HAS2-AS1启动子的组成活性，并鉴定了转录因子结合序列基序。通过siRNA敲低Sp1/Sp3表达可以减弱IL-1 β对HAS2- as1和HAS2的诱导，敲低Smad2/Smad3同样可以减弱tgf - β 1的刺激。使用HAS2- as1特异性sirna抑制IL-1 β刺激的HAS2- as1 RNA诱导也抑制了HAS2 mRNA转录的上调。用硅证明了HAS2- as1 /HAS2异源二聚体形成的热力学可行性，并在体外检测了位点特异性细胞质双链RNA。总之，我们的数据表明，HAS2- as1和HAS2的转录诱导在ptc中同时发生，并且表明反义RNA的转录通过RNA/mRNA异双工形成稳定或增加了这些细胞中HAS2 mRNA的表达。
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-1 beta or TGF-beta 1 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-1 beta induction of both HAS2-AS1 and HAS2, and Smad2/Smad3 knockdown similarly attenuated TGF-beta 1 stimulation. Inhibition of IL-1 beta-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.