The mechanism of transcriptional repression by the nuclear hormone receptor in the preceuse of hormones by using vitamin D receptor as a model system

通过将PTHrP基因中的维生素负反应元件（nVDRE）引入培养细胞，我们获得了以下结果：1)染色质免疫沉淀（ChIP）实验显示，在转染和内源性PTHrP基因中，维生素D将VDR（维生素D受体）和HDAC2（组蛋白去乙酰化酶2）募集到含有nvdrer的染色质上。2) HDAC1、乙酰化组蛋白3 （AcH3）和乙酰化组蛋白4 （AcH4）只有在缺乏维生素D的情况下才会被招募到nVDRE区域。这些结果以及我们使用HDAC抑制剂的实验表明，维生素D通过使nVDRE区域周围的染色质去乙酰化来抑制含有nVDRE基因的表达。3)在随后的免疫沉淀实验中，我们发现VDR以维生素d依赖的方式与HDAC2相互作用。4)此外，通过类似的ChIP实验，我们还发现雌激素将雌激素受体和HDAC2募集到相同的含有nvdrer的染色质区域。同样，HDAC1、AcH3和AcH4仅在没有雌激素的情况下被募集。这些结果表明，配体核激素受体存在一种共同的负基因调控机制。

By introducing the negative vitamin response element (nVDRE) in the PTHrP gene into the cultured cells, we obtained the following results. 1) Chromatin immunoprecipitation (ChIP) assay revealed that vitamin D recruited VDR (vitamin D receptor) and HDAC2 (histone deacetylase 2) to the nVDRE-containing chromatin both in the transfected and endogenous PTHrP genes. 2) HDAC1, acetylated histone3 (AcH3) and acetylated histone 4 (AcH4) were recruited to the nVDRE region only in the absence of vitamin D. These results together with our experiments using a HDAC inhibitor suggested vitamin D repressed the expression of the nVDRE-containing gene through deacetylation of the chromatin surrounding the nVDRE region. 3) In the subsequent immunoprecipitation assay, we found that VDR interacted with HDAC2 in a vitamin D-dependent manner. 4) Further, we also found that estrogen recruited estrogen receptor and HDAC2 to the same nVDRE-containing chromatin region by using similar ChIP assay. Likewise, HDAC1, AcH3 and AcH4 were recruited only in the absence of estrogen. These results suggest that there exists a common machinery for negative gene regulation by the liganded nuclear hormone receptors.