Malone J L, Nelson A C, Lieberman R, Anderson S, Holt J T
Department of Pathology, University of Colorado at Denver (UCDHSC), Aurora, CO 80045, USA.
J Pathol. 2009 Feb;217(3):380-8. doi: 10.1002/path.2458.
Disease-associated BRCA2 mutations typically result in protein truncations that delete the phosphorylation-regulated S3291 BRCA2 domain that interacts with Rad51. BRCA2 hereditary breast cancers are usually ER(+), differing from BRCA1 hereditary cancers, which are usually ER(-). We studied BRCA2 protein expression and S3291 phosphorylation in normal breast tissues and in sporadic breast cancers and observed that BRCA2 is expressed and phosphorylated in normal breast and 10 ER(+) breast cancers but not in 10 ER(-) breast cancers. In order to study this correlation between ER and BRCA2 expression, we studied ER(+) breast cancer cell lines. We found that a rapid increase in BRCA2 S3291 phosphorylation occurs following 17-beta-oestradiol (E2) treatment. This increase seen in BRCA2 total and phospho-S3291 protein levels was found to be unaffected with cycloheximide pre-treatment, but decreased following tamoxifen, ICI 182,780 or roscovitine treatment. This suggests a requirement for ER and cdk (cyclin-dependent kinase) in mediating the increased protein levels. MCF7 cell cycle distribution analysis following E2, in both the presence and absence of roscovitine (a cdk inhibitor), did not demonstrate any changes during an 8 h period, which further supports our hypothesis that mitogenic effects of E2 are not predominant at early time points. Studies with MG132 proteasome inhibitor and siRNA to skp2 support a model in which skp2-mediated proteasomal degradation of BRCA2 rapidly degrades BRCA2 protein in the absence of hormone treatment, which likely inhibits this pathway. E2 was shown to improve survival of MCF7 cells upon radiation treatment and roscovitine partially reversed this effect. We have demonstrated that BRCA2 protein is specifically expressed in ER(+) breast cancers and are investigating a pathway that may show a link between E2 action and BRCA2 protein function in breast cancer.
与疾病相关的BRCA2突变通常导致蛋白质截短，从而缺失与Rad51相互作用的磷酸化调节的S3291 BRCA2结构域。BRCA2遗传性乳腺癌通常为雌激素受体（ER）阳性，这与BRCA1遗传性癌症不同，后者通常为ER阴性。我们研究了正常乳腺组织和散发性乳腺癌中BRCA2蛋白表达及S3291磷酸化情况，发现BRCA2在正常乳腺和10例ER阳性乳腺癌中表达且发生磷酸化，但在10例ER阴性乳腺癌中未出现这种情况。为了研究ER与BRCA2表达之间的这种相关性，我们对ER阳性乳腺癌细胞系进行了研究。我们发现，17-β-雌二醇（E2）处理后，BRCA2 S3291磷酸化迅速增加。BRCA2总蛋白水平和磷酸化S3291蛋白水平的这种增加不受放线菌酮预处理的影响，但在他莫昔芬、ICI 182,780或罗哌卡因处理后降低。这表明介导蛋白水平增加需要ER和细胞周期蛋白依赖性激酶（cdk）。在有或没有罗哌卡因（一种cdk抑制剂）的情况下，E2处理后MCF7细胞周期分布分析在8小时内未显示任何变化，这进一步支持了我们的假设，即E2的促有丝分裂作用在早期时间点并不占主导地位。使用MG132蛋白酶体抑制剂和针对skp2的小干扰RNA（siRNA）的研究支持了一种模型，即在没有激素处理的情况下，skp2介导的BRCA2蛋白酶体降解会迅速降解BRCA2蛋白，这可能会抑制该途径。E2被证明可提高MCF7细胞在放疗后的存活率，而罗哌卡因可部分逆转这种作用。我们已经证明BRCA2蛋白在ER阳性乳腺癌中特异性表达，并且正在研究一条可能显示E2作用与乳腺癌中BRCA2蛋白功能之间联系的途径。
