Heterozygosity for mutations in the BRCA1 gene in humans confers high risk for developing breast cancer, but a biochemical basis for this phenotype has not yet been determined. Evidence has accumulated implicating BRCA1, in the maintenance of genomic integrity and the protection of cells against DNA double strand breaks (DSB). Here we present evidence that human cells heterozygous for BRCA1 mutations exhibit impaired DNA end-joining, which is the major DSB repair pathway in mammalian somatic cells. Using an in vivo host cell end-joining assay, we observed that the ®delity of DNA end-joining is strongly reduced in three BRCA1 +/7 cell lines in comparison to two control cell lines. Moreover, cell-free BRCA1 +/7 extracts are unable to promote accurate DNA end-joining in an in vitro reaction. The steady-state level of the wild type BRCA1 protein was signi®cantly lower than the 50% expected in BRCA1 +/7 cells and thus may underlie the observed endjoining defect. Together, these data strongly suggest that BRCA1 is necessary for faithful rejoining of broken DNA ends and that a single mutated BRCA1 allele is sucient to impair this process. This defect will compromise genomic stability in BRCA1 germ-line mutation carriers, triggering the genetic changes necessary for the initiation of neoplastic transformation.

DNA end-joining is the major repair pathway for double-strand breaks (DSBs) in higher eukaryotes. To understand how DSB structure affects the end-joining process in human cells, we have examined the in vivo repair of linearized plasmids containing complementary as well as several different configurations of non-complementary DNA ends. Our results demonstrate that, while complementary and blunt termini display comparable levels of error-free rejoining, end-joining fidelity is decreased to varying extents among mismatched non-complementary ends. End structure also influences the kinetics of repair, accurately recircularized substrates for blunt and complementary termini being detected significantly earlier than for mismatched non-complementary ends. These results suggest that the end-joining process is composed of an early component, capable of efficiently repairing substrates requiring a single ligation event, and a late component, involved in the rejoining of complex substrates requiring multiple processing steps. Finally, these two types of repair events may have different genetic requirements as suggested by the finding that exposure of cells to wortmannin, a potent inhibitor of phosphatidylinositol 3-related kinases (PI 3-related kinases), blocks the repair of complex substrates while having little or no effect on those requiring a simple ligation event.

This preliminary report suggests an association, not previously reported, between maternal HIV-positive status and an increased risk of necrotizing enterocolitis in premature infants. Despite the limitations of this study, we suggest that premature newborn infants of HIV-positive mothers should be monitored very carefully for a possible increased risk of necrotizing enterocolitis.