The gene encoding p53 mediates a major tumor suppression pathway that is frequently altered in human cancers. p53 function is kept at a low level during normal cell growth and is activated in response to various cellular stresses. The MDM2 oncoprotein plays a key role in negatively regulating p53 activity by either direct repression of p53 transactivation activity in the nucleus or promotion of p53 degradation in the cytoplasm. DNA damage and oncogenic insults, the two best-characterized p53-dependent checkpoint pathways, both activate p53 through inhibition of MDM2. Here we report that the human homologue of MDM2, HDM2, binds to ribosomal protein L11. L11 binds a central region in HDM2 that is distinct from the ARF binding site. We show that the functional consequence of L11-HDM2 association, like that with ARF, results in the prevention of HDM2-mediated p53 ubiquitination and degradation, subsequently restoring p53-mediated transactivation, accumulating p21 protein levels, and inducing a p53-dependent cell cycle arrest by canceling the inhibitory function of HDM2. Interference with ribosomal biogenesis by a low concentration of actinomycin D is associated with an increased L11-HDM2 interaction and subsequent p53 stabilization. We suggest that L11 functions as a negative regulator of HDM2 and that there might exist in vivo an L11-HDM2-p53 pathway for monitoring ribosomal integrity.

The p53-inhibitory function of the oncoprotein MDM2 is regulated by a number of MDM2-binding proteins, including ARF and ribosomal proteins L5, L11, and L23, which bind the central acidic domain of MDM2 and inhibit its E3 ubiquitin ligase activity. Various human cancer-associated MDM2 alterations targeting the central acidic domain have been reported, yet the functional significance of these mutations in tumor development has remained unclear. Here, we show that cancer-associated missense mutations targeting MDM2's central zinc finger disrupt the interaction of MDM2 with L5 and L11. We found that the zinc finger mutant MDM2 is impaired in undergoing nuclear export and proteasomal degradation as well as in promoting p53 degradation, yet retains the function of suppressing p53 transcriptional activity. Unlike the wild-type MDM2, whose p53-suppressive activity can be inhibited by L11, the MDM2 zinc finger mutant escapes L11 inhibition. Hence, the MDM2 central zinc finger plays a critical role in mediating MDM2's interaction with ribosomal proteins and its ability to degrade p53, and these roles are disrupted by human cancer-associated MDM2 mutations.The mammalian p53 transcription factor mediates a major tumor suppression pathway that is negatively controlled by the proto-oncoprotein MDM2 (HDM2 in humans; henceforth denoted MDM2) and is altered in most, if not all, human cancers. The gene for mouse Mdm2 (murine double minute 2) was originally identified in a spontaneously transformed mouse BALB/c cell line (13). The Mdm2 protein was found to be responsible for transformation of NIH 3T3 and Rat2 cells when overexpressed (13), and this transforming function is believed to stem from its ability to bind with and inhibit the transactivation activity of p53 (39). Subsequently, the HDM2 gene, the human homologue of Mdm2, was found to be amplified in over one-third of those human sarcomas that still retain wild-type p53 (41), suggesting that overexpression of MDM2 could be a common mechanism by which cells inactivate p53. Mice with targeted deletion of the Mdm2 gene die during early embryonic development, and this lethality can be rescued by concomitant deletion of p53, indicating that a major in vivo function of MDM2 is to keep p53 activity in check (27,33).It is believed that MDM2 controls p53 through two mechanisms: inhibition of the transcriptional activity of p53 (39) and promotion of p53 ubiquitination and degradation (18,29). Mdm2 binds to and masks the N-terminal transactivation domain of p53 by directly interfering with the interaction between p53 and the basal transcriptional machinery (42, 54). Mdm2 belongs to a large family of RING finger ubiquitin ligases (25). Studies have demonstrated that Mdm2 is a ubiquitin ligase (19) and that the ubiquitin ligase activity of Mdm2 is responsible for degradation of p53 both in vitro (14,20) and in transfected cells (14). MDM2-mediated p53 degradation also depends on its ability to promote p53 nuclear export (46). Mutation of a nuclear export signal (NES) in MDM2 abolishes ...