RBP1 Recruits the mSIN3-Histone Deacetylase Complex to the Pocket of Retinoblastoma Tumor Suppressor Family Proteins Found in Limited Discrete Regions of the Nucleus at Growth Arrest

Retinoblastoma (RB) tumor suppressor family pocket proteins induce cell cycle arrest by repressing transcription of E2F-regulated genes through both histone deacetylase (HDAC)-dependent and -independent mechanisms. In this study we have identified a stable complex that accounts for the recruitment of both repression activities to the pocket. One component of this complex is RBP1, a known pocket-binding protein that exhibits both HDAC-dependent and -independent repression functions. RB family proteins were shown to associate via the pocket with previously identified mSIN3-SAP30-HDAC complexes containing exclusively class I HDACs. Such enzymes do not interact directly with RB family proteins but rather utilize RBP1 to target the pocket. This mechanism was shown to account for the majority of RB-associated HDAC activity. We also show that in quiescent normal human cells this entire RBP1-mSIN3-SAP30-HDAC complex colocalizes with both RB family members and E2F4 in a limited number of discrete regions of the nucleus that in other studies have been shown to represent the initial origins of DNA replication following growth stimulation. These results suggest that RB family members, at least in part, drive exit from the cell cycle by recruitment of this HDAC complex via RBP1 to repress transcription from E2F-dependent promoters and possibly to alter chromatin structure at DNA origins.

视网膜母细胞瘤（Retinoblastoma, RB）抑癌蛋白家族口袋蛋白可通过组蛋白去乙酰化酶（histone deacetylase, HDAC）依赖与非依赖两种机制抑制E2F调控基因的转录，从而诱导细胞周期阻滞。本研究鉴定出一种可将两种抑制活性募集至口袋结构域的稳定复合物。该复合物的组分之一为RBP1，这是一种已被证实兼具HDAC依赖与非依赖抑制功能的已知口袋结合蛋白。研究表明，RB家族蛋白可通过口袋结构域与此前已鉴定的仅含I类HDAC的mSIN3-SAP30-HDAC复合物结合。这类酶并不会直接与RB家族蛋白相互作用，而是通过RBP1靶向口袋结构域，该机制可解释绝大多数RB相关的HDAC活性。本研究同时证实，在静息正常人类细胞中，整个RBP1-mSIN3-SAP30-HDAC复合物可与RB家族成员及E2F4共定位于细胞核内有限数量的离散区域；既往研究显示，这些区域正是生长刺激后DNA复制的初始起始位点。上述结果提示，RB家族蛋白至少可部分通过RBP1募集该HDAC复合物，以抑制E2F依赖型启动子的转录，并可能改变DNA复制起始位点处的染色质结构，从而驱动细胞周期退出。