Members of the enhancer of split- and hairy-related protein (SHARP) family, SHARP-1 and SHARP-2, are basic helix−loop−helix transcriptional repressors and belong to the clock genes. In this study, an effect of retinoic acid (RA) on the SHARP family gene expression in the differentiated cells was examined. RA rapidly and temporarily induced the SHARP-2 mRNA expression in hepatic H4IIE cells. Then, whether the SHARP-2 mRNA expression is altered by dexamethasone (Dex), insulin, and the combination of RA and Dex or RA and insulin was examined. Dex had different effects on the expression of SHARP-2 mRNA in the presence or absence of RA. Then, the molecular mechanisms were investigated using inhibitors of various signaling molecules. The RA-induction of SHARP-2 mRNA level was mainly inhibited by LY294002, staurosporine, and actinomycin D, respectively. Finally, whether RA acts on the transcriptional regulatory region of the SHARP-2 gene was analysed using luciferase reporter gene assay. At least two RA-responsive regions were mapped at the nucleotide sequences between −3,700 and −1,600 of the SHARP-2 gene. In addition, this effect was dependent on the RA receptor and retinoid X receptor. Thus, we conclude that RA stimulated transcription of the SHARP-2 gene via multiple pathways.

中文翻译：

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维甲酸通过多种途径刺激大鼠 SHARP-2 基因的转录

分裂和多毛相关蛋白 (SHARP) 家族增强子 SHARP-1 和 SHARP-2 的成员是基本的螺旋-环-螺旋转录抑制因子，属于时钟基因。在这项研究中，检查了视黄酸 (RA) 对分化细胞中 SHARP 家族基因表达的影响。RA 在肝 H4IIE 细胞中迅速和暂时地诱导 SHARP-2 mRNA 表达。然后，检查地塞米松(Dex)、胰岛素以及RA和Dex或RA和胰岛素的组合是否改变了SHARP-2 mRNA表达。在存在或不存在 RA 的情况下，Dex 对 SHARP-2 mRNA 的表达有不同的影响。然后，使用各种信号分子的抑制剂研究分子机制。SHARP-2 mRNA 水平的 RA 诱导主要被 LY294002、星形孢菌素和放线菌素 D 分别抑制。最后，使用荧光素酶报告基因测定法分析 RA 是否作用于 SHARP-2 基因的转录调控区。在 SHARP-2 基因的 -3,700 和 -1,600 之间的核苷酸序列上绘制了至少两个 RA 反应区域。此外，这种作用依赖于 RA 受体和类视黄醇 X 受体。因此，我们得出结论，RA 通过多种途径刺激了 SHARP-2 基因的转录。