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DHRS4/NRDR基因编码一种属于SDR家族的酶,在维甲酸合成、类固醇代谢和苯甲基代谢中发挥生物合成催化作用.DHRS4基因定位于14q11.2,有两个相似的拷贝基因,分别为DHRS4L2和DHRS4L1.我们前期发现了DHRS4L2基因一个上游转录起始位点,命名为DHRS4L2-Ea.在本研究中,我们用RT-PCR和双脱氧测序法发现一个新的从DHRS4L2-Ea转录的选择性剪接亚型DHRS4L2-900a(KC237374).同时RT-PCR结果显示在SK-N-SH细胞DHRS4L2-Ea选择性剪接亚型中DHRS4L2-iso(AY616183)表达最多,为主要亚型.在SK-N-SH细胞过表达DHRS4L2-800a(AY920361)使DHRS4L2-Ea基因下游CPNE6 mRNA表达下调.在HeLa细胞过表达DHRS4L2-800a(AY920361)或DHRS4L2-900a(KC237374)进一步表明DHRS4L2-Ea抑制CPNE6表达的作用.定量PCR结果显示si-RNA抑制DHRS4L2-Ea表达使CPNE6 mRNA表达上调.亚硫酸盐测序结果显示在SK-N-SH转染DHRS4L2-800a(AY920361)的样本中CPNE6基因DNA CpG甲基化增加.综上所述,本研究揭示DHRS4L2表达的非编码RNA抑制其下游基因CPNE6的表达.
The DHRS4 / NRDR gene encodes an enzyme belonging to the SDR family and plays a biosynthetic catalytic role in retinoic acid synthesis, steroid metabolism and benzyl metabolism. The DHRS4 gene is located at 14q11.2 and has two similar copy genes, namely DHRS4L2 and DHRS4L1. We found an upstream transcriptional start site of DHRS4L2 gene and named DHRS4L2-Ea.In this study, we found a new transcription from DHRS4L2-Ea by RT-PCR and dideoxy sequencing DHRS4L2-900a (KC237374) .At the same time, RT-PCR results showed that DHRS4L2-iso (AY616183) was the most expressed and predominant DHRS4L2-Ea alternative splicing subtype in SK-N-SH cells, Overexpression of DHRS4L2-800a (AY920361) in N-SH cells downregulated the expression of CPNE6 mRNA downstream of DHRS4L2-Ea gene.Expression of DHRS4L2-800a (AY920361) or DHRS4L2-900a (KC237374) further demonstrated that DHRS4L2-Ea inhibited CPNE6 expression in HeLa cells The results of quantitative PCR showed that siRNA inhibited the expression of CPE6 mRNA by upregulating the expression of DHRS4L2-Ea.The results of sulphite sequencing showed that DNA methylation of CPNE6 was detected in the samples of SK-N-SH transfected DHRS4L2-800a (AY920361) In summary, the study reveals Expression of non-coding RNA which inhibits expression of the downstream gene DHRS4L2 of CPNE6.