Inactivation of cell death is a major step in tumor development, and p53, a tumor suppressor frequently mutated in cancer, is a critical mediator of cell death. While a role for p53 in apoptosis is well established, direct links to other pathways controlling cell death are unknown. Here we describe DRAM (damage-regulated autophagy modulator), a p53 target gene encoding a lysosomal protein that induces macroautophagy, as an effector of p53-mediated death. We show that p53 induces autophagy in a DRAM-dependent manner and, while overexpression of DRAM alone causes minimal cell death, DRAM is essential for p53-mediated apoptosis. Moreover, analysis of DRAM in primary tumors revealed frequent decreased expression often accompanied by retention of wild-type p53. Collectively therefore, these studies not only report a stress-induced regulator of autophagy but also highlight the relationship of DRAM and autophagy to p53 function and damage-induced programmed cell death.

The status of p53 was investigated in breast tumours arising in germ-line carriers of mutant alleles of BRCA1 and BRCA2 and in a control series of sporadic breast tumours. p53 expression was detected in 20/26 (77%) BRCA1-, 10/22 (45%) BRCA2-associated and 25/72 (35%) grade-matched sporadic tumours. Analysis of p53 sequence revealed that the gene was mutant in 33/50 (66%) BRCA-associated tumours, whereas 7/20 (35%) sporadic grade-matched tumours contained p53 mutation (P50.05). A number of the mutations detected in the BRCA-associated tumours have not been previously described in human cancer databases, whilst others occur extremely rarely. Analysis of additional genes, p16 INK4 , Ki-ras and b-globin revealed absence or very low incidence of mutations, suggesting that the higher frequency of p53 mutation in the BRCA-associated tumours does not re¯ect a generalized increase in susceptibility to the acquisition of somatic mutation. Furthermore, absence of frameshift mutations in the polypurine tracts present in the coding sequence of the TGF b type II receptor (TGF b IIR) and Bax implies that loss of function of BRCA1 or BRCA2 does not confer a mutator phenotype such as that found in tumours with microsatellite instability (MSI). p21 Waf1 was expressed in BRCA-associated tumours regardless of p53 status and, furthermore, some tumours expressing wild-type p53 did not express detectable p21 Waf1 . These data do not support, therefore, the simple model based on studies of BRCA7/7 embryos, in which mutation of p53 in BRCA-associated tumours results in loss of p21 Waf1 expression and deregulated proliferation. Rather, they imply that proliferation of such tumours will be subject to multiple mechanisms of growth regulation.

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Inheritance of germ-line mutant alleles of BRCA1 and BRCA2 confers a markedly increased risk of breast cancer and we have previously reported a higher incidence of p53 mutations in these tumours than in grade matched sporadic tumours. We have now characterized these p53 mutants. The results of these studies identify a novel class of p53 mutants previously undescribed in human cancer yet with multiple occurrences in BRCA-associated tumours which retain a pro®le of p53-dependent activities in terms of transactivation, growth suppression and apoptosis induction which is close or equal to wild-type. However, these mutants fail to suppress transformation and exhibit gain of function transforming activity in rat embryo ®broblasts. These mutants therefore fall into a novel category of p53 mutants which dissociate transformation suppression from other wild-type functions. The rarity of these mutants in human cancer and their multiple occurrence in BRCA-associated breast tumours suggests that these novel p53 mutants are selected during malignant progression in the unique genetic background of BRCA1-and BRCA2-associated tumours.