Oestrogen-mediated phosphorylation and stabilization of BRCA2 protein in breast.
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.
Malone
J L
JL
Department of Pathology, University of Colorado at Denver (UCDHSC), Aurora, CO 80045, USA.
Nelson
A C
AC
Lieberman
R
R
Anderson
S
S
Holt
J T
JT
D024682:BRCA2 Protein / Q000032:analysis / Q000378:metabolism*; D015153:Blotting, Western / Q000379:methods; D001940:Breast / Q000378:metabolism; D001943:Breast Neoplasms / Q000378:metabolism*; D045744:Cell Line, Tumor; D004260:DNA Repair; D004967:Estrogens / Q000378:metabolism / Q000494:pharmacology*; D005260:Female; D006801:Humans; D007150:Immunohistochemistry; D010766:Phosphorylation; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods
D024682:BRCA2 Protein; D004967:Estrogens