Individuals carrying mutations in BRCA1 or p53 genes are predisposed to a variety of cancers, and both tumor suppressor genes have been implicated in DNA damage response pathways. We have analyzed a possible functional link between p53 and BRCA1 genes. Here we show that BRCA1 expression levels are down-regulated in response to p53 induction in cells that undergo either growth arrest, senescence, or apoptosis. Physiological stimuli, such as exposure to DNA-damaging agents, also result in negative regulation of BRCA1 levels in a p53-dependent manner prior to causing cell cycle arrest. Nuclear run-on experiments and luciferase reporter assays demonstrate that the changes in BRCA1 expression are mainly due to transcriptional repression induced by p53. In conclusion, the data show that BRCA1 expression levels are controlled by the presence and activity of wild-type p53 and suggest the existence of an intracellular p53/BRCA1 pathway in the response of cells to stress conditions. p53, the protein product of a tumor suppressor gene, has been implicated in the control of cell proliferation and tumor progression, as well as in the maintenance of genome integrity in response to DNA-damaging events (1,28,29). Deletion or inactivation of the p53 gene is observed in more than half of human cancers (23). p53 is induced in response to suboptimal growth conditions (DNA damage, hypoxia, heat, starvation, etc.) and acts as an "emergency brake" to trigger cells to reversible or irreversible growth arrest or even to apoptosis (8,15,20,38,54,62), thus protecting the genome from accumulating an excess of mutations. Once active, p53 binds specifically to p53 response elements and transactivates the expression of genes such as those encoding mdm2, GADD45, p21 (Waf1 or Cip1), bax, PAG608, cyclin G, and IGF-BP3 (1,26,28,29). The p53 protein has also been shown to repress transcription driven from certain viral promoters, as well as from cellular genes. Genes encoding RB, c-fos, MAP4, presenilin, and DNA topoisomerase II␣ are among the characterized p53-repressed genes (13,38,42,50,58).Given the complexity and importance of the cellular response to p53, its expression must be tightly regulated. The ubiquitin-mediated proteolysis pathway is, at least in part, responsible for maintaining p53 protein levels at a low concentration in normal cells (9). p53 levels and activity are also under tight control by upstream effectors such as the DNAprotein kinase family (60), negative autoregulation (34), phosphorylation (24,25,55,59), and other cellular proteins such as mdm2 and CBP/p300 (17,30,32,49,57).BRCA1 is also a tumor suppressor gene. Mutations in the BRCA1 gene account for about 50% of inherited breast cancer cases and 80% of families predisposed to breast and ovarian cancers (5, 39). In fact, there are interesting parallels between p53 and BRCA1. BRCA1, like p53, is a cell cycle-regulated nuclear phosphoprotein (27,43,56) and has also been implicated in DNA damage response and repair pathways. In addition, BRCA1 interacts with some of the ma...

Two 29 kDa subunits of the multicatalytic proteinase (proteasome) complex, the C8 and C9 components, are phosphorylated in vivo and can be phosphorylated in vitro by casein kinase II (CKII). The major phosphate acceptor is the C8 subunit being phosphorylated in serine, both in vivo and in vitro. The phosphopeptides generated by Glu-C endoprotease digestion from the in vivo 29 kDa labeled subunit and from the in vitro phosphorylation of the recombinant C8 subunit with CKII are identical, suggesting that CKII is likely responsible for the in vivo phosphorylation of the C8 subunit. The in vitro stoichiometry of phosphorylation of the proteasome complex and the recombinant C9 and C8 subunits by CKII is 2-2.5, 0.2, and 2 mol of phosphate per mole, respectively. Several C8 protein constructs allow the location of the CKII phosphorylation sites to be the COOH terminal portion of the protein, and direct mutational analyses show that Ser-243 and Ser-250 are the residues of the C8 subunit phosphorylated by CKII. The in vitro phosphorylation of the proteasome by CKII does not affect its proteolytic activity (on proteins or fluorogenic synthetic peptides), therefore suggesting its involvement in the interaction of the proteasome with other cellular proteins, i.e. in the formation of the 26S complex and/or in the interaction with the nuclear translocation machinery.