Disruption of BRCA1 LXCXE motif alters BRCA1 functional activity and regulation of RB family but not RB protein binding
Disruption of BRCA1 LXCXE motif alters BRCA1 functional activity and regulation of RB family but not RB protein binding
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DOI：
10.1038/sj.onc.1204666
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发表时间：
2001-08-09
期刊：
影响因子：
8
通讯作者：
Rosen, EM
中科院分区：
文献类型：
--
作者：
Fan, SJ;Yuan, RQ;Rosen, EM
BRCA1 基因产物的肿瘤抑制活性部分归因于与其他肿瘤抑制因子（包括 p53 和视网膜母细胞瘤 (RB) 蛋白）的功能相互作用。 BRCA1 上的 RB 结合位点在 C 端 BRCT 结构域 (Yarden 和 Brody, 1999) 和 N 端 (aa 304-394) (Aprelikova 等人, 1999) 中被鉴定。 N 端位点包含共有 RB 结合基序 LXCXE (aa 358-362)，但该基序在 RB 结合和 BRCA1 功能活性中的作用尚不清楚。在体外和体内测定中，我们发现 BRCA1:RB 相互作用不需要 BRCA1 LXCXE 基序，也不需要 RB 的完整 A/B 结合袋。此外，还观察到内源性 BRCA1 和 RB 蛋白的核共定位。野生型 BRCA1 (wtBRCA1) 的过度表达不会导致细胞周期停滞，但会导致 RB、p107、p130 和其他蛋白（例如 p300）表达下调，与对 DNA 损伤剂的敏感性增加相关。相反，具有LXCXE失活突变（LXCXE-->RXRXH）的全长BRCA1的表达未能下调RB，阻断wtBRCA1对RB的下调，诱导化疗耐药性，并消除BRCA1介导DU-145前列腺癌细胞肿瘤生长抑制的能力。 wtBRCA1 诱导的化疗敏感性可通过 Rb 或 p300 的表达部分逆转，并通过 Rb 加 p300 的共表达完全逆转。我们的研究结果表明：（I）N 端 RB 结合区域内 LXCXE 基序的破坏改变了 BRCA1 的生物学功能； (2)BRCA1的过度表达抑制RB和RB家族(p107和p130)蛋白的表达。
The tumor suppressor activity of the BRCA1 gene product is due, in part, to functional interactions with other tumor suppressors, including p53 and the retinoblastoma (RB) protein. RB binding sites on BRCA1 were identified in the C-terminal BRCT domain (Yarden and Brody, 1999) and in the N-terminus (aa 304-394) (Aprelikova et al., 1999). The N-terminal site contains a consensus RB binding motif, LXCXE (aa 358-362), but the role of this motif in RB binding and BRCA1 functional activity is unclear. In both in vitro and in vivo assays, we found that the BRCA1:RB interaction does not require the BRCA1 LXCXE motif, nor does it require an intact A/B binding pocket of RB. In addition, nuclear co-localization of the endogenous BRCA1 and RB proteins was observed. Over-expression of wild-type BRCA1 (wtBRCA1) did not cause cell cycle arrest but did cause down-regulation of expression of RB, p107, p130, and other proteins (e.g., p300), associated with increased sensitivity to DNA-damaging agents. In contrast, expression of a full-length BRCA1 with an LXCXE inactivating mutation (LXCXE-->RXRXH) failed to down-regulate RB, blocked the down-regulation of RB by wtBRCA1, induced chemoresistance, and abrogated the ability of BRCA1 to mediate tumor growth suppression of DU-145 prostate cancer cells. wtBRCA1-induced chemosensitivity was partially reversed by expression of either Rb or p300 and fully reversed by co-expression of Rb plus p300. Our findings suggest that: (I) disruption of the LXCXE motif within the N-terminal RB binding region alters the biologic function of BRCA1; and (2) over-expression of BRCA1 inhibits the expression of RB and RB family (p107 and p130) proteins.