Impaired DNA Damage Response in Cells Expressing an Exon 11-Deleted Murine Brca1 Variant That Localizes to Nuclear Foci

Abstract

Both human and mouse cells express an alternatively spliced variant of BRCA1, BRCA1-Δ11, which lacks exon 11 in its entirety, including putative nuclear localization signals. Consistent with this, BRCA1-Δ11 has been reported to reside in the cytoplasm, a localization that would ostensibly preclude it from playing a role in the nuclear processes in which its full-length counterpart has been implicated. Nevertheless, the finding that murine embryos bearing homozygous deletions of exon 11 survive longer than embryos that are homozygous for Brca1 null alleles suggests that exon 11-deleted isoforms may perform at least some of the functions of Brca1. We have analyzed both the full-length and the exon 11-deleted isoforms of the murine Brca1 protein. Our results demonstrate that full-length murine Brca1 is identical to human BRCA1 with respect to its cell cycle regulation, DNA damage-induced phosphorylation, nuclear localization, and association with Rad51. Surprisingly, we show that endogenous Brca1-Δ11 localizes to discrete nuclear foci indistinguishable from those found in wild-type cells, despite the fact that Brca1-Δ11 lacks previously defined nuclear localization signals. However, we further show that DNA damage-induced phosphorylation of Brca1-Δ11 is significantly reduced compared to full-length Brca1, and that gamma irradiation-induced Rad51 focus formation is impaired in cells in which only Brca1-Δ11 is expressed. Our results suggest that the increased viability of embryos bearing homozygous deletions of exon 11 may be due to expression of Brca1-Δ11 and suggest an explanation for the genomic instability that accompanies the loss of full-length Brca1.

ACKNOWLEDGMENTS

We thank members of the Chodosh laboratory and Prakash K. Rao for helpful discussions and critical reading of the manuscript. Confocal microscopy was made possible by the help of James F. Sanzo and Irina Chernysh of the Biomedical Imaging Core Facility at the University of Pennsylvania Medical Center.

This research was supported by NIH grants CA71513 and CA78410 from the National Cancer Institute, and U. S. Army Breast Cancer Research Program grants DAMD17-98-1-8230 (L.J.H.), DAMD17-96-1-6111 (S.R.M.), DAMD17-00-1-0403 (C.J.S.), DAMD17-98-1-8226, and DAMD17-96-1-6113.