Pemsel Anja, Rumpf Saskia, Roemer Klaus, Heyne Kristina, Vogt Thomas, Reichrath Joerg
Department of Dermatology, The Saarland University Hospital, Homburg, Germany.
José Carreras Center and Internal Medicine I, University of Saarland Medical Center, Homburg, Germany.
Anticancer Res. 2018 Feb;38(2):1209-1216. doi: 10.21873/anticanres.12341.
While nuclear cofactors that contribute to vitamin D receptor (VDR)-mediated gene transcription, including retinoid X receptors, nuclear co-activators and co-repressors, have been extensively investigated, little is known about cytoplasmic VDR-binding partners and the physiological relevance of their interaction. To gain new insight into this topic, we isolated whole-cell protein extracts of 1,25-dihydroxyvitamin D stimulated and UV-B-irradiated vs. non-irradiated HEK 293T cells transfected with a plasmid called pURB VDR C-Term TAP tag. VDR complex was purified by tandem affinity purification (TAP). The nuclear tumor-suppressor protein p53 and its negative regulator novel INHAT repressor (NIR), in addition to 43 other nuclear or cytoplasmatic VDR binding partners, were identified using nano high-performance liquid chromatography-electrospray ionization tandem mass spectrometric analysis. VDR binding to p53 was confirmed by western blot analysis. Future studies are required to further elucidate the functional significance of these interactions.
虽然对有助于维生素D受体（VDR）介导的基因转录的核辅因子，包括视黄酸X受体、核共激活因子和共抑制因子，已经进行了广泛研究，但对于细胞质VDR结合伴侣及其相互作用的生理相关性却知之甚少。为了深入了解这一主题，我们分离了用名为pURB VDR C-Term TAP标签的质粒转染的、经1,25 - 二羟基维生素D刺激和UV - B照射与未照射的HEK 293T细胞的全细胞蛋白提取物。通过串联亲和纯化（TAP）纯化VDR复合物。使用纳米高效液相色谱 - 电喷雾电离串联质谱分析鉴定了核肿瘤抑制蛋白p53及其负调节因子新型INHAT抑制因子（NIR），以及其他43种核或细胞质VDR结合伴侣。通过蛋白质印迹分析证实了VDR与p53的结合。需要进一步的研究来阐明这些相互作用的功能意义。
