Nuclear coactivator-62 kDa/Ski-interacting protein is a nuclear matrix-associated coactivator that may couple vitamin D receptor-mediated transcription and RNA splicing
Nuclear coactivator-62 kDa/Ski-interacting protein is a nuclear matrix-associated coactivator that may couple vitamin D receptor-mediated transcription and RNA splicing
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DOI：
10.1074/jbc.m305191200
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发表时间：
2003-09-12
影响因子：
4.8
通讯作者：
MacDonald, PN
中科院分区：
文献类型：
--
作者：
Zhang, C;Dowd, DR;MacDonald, PN
核辅激活因子-62 kDa/Ski-interacting protein（NCoA 62/SKIP）是一种推定的维生素D受体（VDR）和核受体辅激活因子蛋白，与其他VDR辅激活因子无关，如类固醇受体辅激活因子（SRC）家族中的那些。NCoA 62/SKIP在VDR激活的转录中发挥作用的机制尚不清楚。在本研究中，我们确定了NCoA 62/SKIP的COOH末端的核定位序列，并表明NCoA 62/SKIP靶向核基质亚结构域。染色质免疫沉淀研究显示，内源性NCoA 62/SKIP以1，25-二羟维生素D-3依赖的方式与ROS 17/2.8骨肉瘤细胞中的VDR靶基因相关。观察到VDR、SRC-1和NCoA 62/SKIP占据启动子的循环模式，NCoA 62/SKIP在SRC-1之后进入这些启动子复合物。这些研究为NCoA 62/SKIP作为VDR转录共激活因子的作用提供了强有力的支持，并且它们表明NCoA 62/SKIP和SRC共激活因子之间可能存在关键的机制差异。为了探索潜在的机制，NCoA 62/SKIP相互作用蛋白从HeLa细胞核提取物中纯化，并通过质谱鉴定。所鉴定的蛋白质代表剪接体以及其他核基质相关蛋白的组分。在这里，我们表明，显性负抑制剂NCoA 62/SKIP（dnNCoA 62/SKIP）干扰适当剪接的转录衍生自1，25-二羟维生素D-3诱导的生长激素minigene盒的表达。总之，这些数据表明，NCoA 62/SKIP具有与核受体共激活因子和RNA剪接体组分一致的性质，从而表明NCoA 62/SKIP在将VDR介导的转录与RNA剪接偶联中的潜在作用。
Nuclear coactivator-62 kDa/Ski-interacting protein (NCoA62/SKIP) is a putative vitamin D receptor (VDR) and nuclear receptor coactivator protein that is unrelated to other VDR coactivators such as those in the steroid receptor coactivator ( SRC) family. The mechanism through which NCoA62/SKIP functions in VDR-activated transcription is unknown. In the present study, we identified a nuclear localization sequence in the COOH terminus of NCoA62/SKIP and showed that NCoA62/SKIP was targeted to nuclear matrix subdomains. Chromatin immunoprecipitation studies revealed that endogenous NCoA62/SKIP associated in a 1,25-dihydroxyvitamin D-3-dependent manner with VDR target genes in ROS17/2.8 osteosarcoma cells. A cyclic pattern of promoter occupancy by VDR, SRC-1, and NCoA62/SKIP was observed, with NCoA62/SKIP entering these promoter complexes after SRC-1. These studies provide strong support for the proposed role of NCoA62/SKIP as a VDR transcriptional coactivator, and they indicate that key mechanistic differences probably exist between NCoA62/SKIP and SRC coactivators. To explore potential mechanisms, NCoA62/SKIP-interacting proteins were purified from HeLa cell nuclear extracts and identified by mass spectrometry. The identified proteins represent components of the spliceosome as well as other nuclear matrix-associated proteins. Here, we show that a dominant negative inhibitor of NCoA62/SKIP (dnNCoA62/SKIP) interfered with appropriate splicing of transcripts derived from 1,25-dihydroxyvitamin D-3-induced expression of a growth hormone minigene cassette. Taken together, these data show that NCoA62/SKIP has properties that are consistent with those of nuclear receptor coactivators and with RNA spliceosome components, thus suggesting a potential role for NCoA62/SKIP in coupling VDR-mediated transcription to RNA splicing.