Inhibition of HDM2 and Activation of p53 by Ribosomal Protein L23

Abstract

The importance of coordinating cell growth with proliferation has been recognized for a long time. The molecular basis of this relationship, however, is poorly understood. Here we show that the ribosomal protein L23 interacts with HDM2. The interaction involves the central acidic domain of HDM2 and an N-terminal domain of L23. L23 and L11, another HDM2-interacting ribosomal protein, can simultaneously yet distinctly interact with HDM2 together to form a ternary complex. We show that, when overexpressed, L23 inhibits HDM2-induced p53 polyubiquitination and degradation and causes a p53-dependent cell cycle arrest. On the other hand, knocking down L23 causes nucleolar stress and triggers translocation of B23 from the nucleolus to the nucleoplasm, leading to stabilization and activation of p53. Our data suggest that cells may maintain a steady-state level of L23 during normal growth; alternating the levels of L23 in response to changing growth conditions could impinge on the HDM2-p53 pathway by interrupting the integrity of the nucleolus.The tumor suppressor protein p53 plays a pivotal role in preventing damaged and abnormal cells from becoming malignant, and its loss of function is associated with a majority of human cancers (26,36,37). The activity of p53 is not required for normal cell growth, and the protein is kept at low levels and inactive. This is accomplished by the proto-oncoprotein HDM2 (human counterpart of MDM2 in mice) through either ubiquitin-dependent p53 degradation in the cytoplasm (6, 7, 11) or repression of p53 transcriptional activity in the nucleus (17, 35). The HDM2 gene can, in turn, be transcriptionally activated by p53, constituting a feedback regulatory loop (1, 40). The current understanding of the major mechanisms of p53 activation includes one that is triggered by DNA damage and induces p53 phosphorylation through a cascade of protein kinases (5, 15, 32) and one that is triggered by aberrant oncogenic signals and is mediated by p14 ARF (p19 ARF in mice) (31). Little is known about the connection between p53 and ribosomal biogenesis, even though ribosomal biogenesis occupies the major part of a cell's life cycle. Whether and how p53 may be involved in checking ribosomal stress and the integrity of ribosomal biogenesis remains elusive (28).Previous studies have identified the interaction of the ribosomal protein L5 with HDM2 (14). Recently, it has been shown that the ribosomal protein L11 also interacts with HDM2, and through this interaction, L11 stabilizes and activates p53 and induces a cell cycle arrest (13,41). This is achieved, at least in part, through L11 inhibiting the E3 ligase activity of HDM2 (41). Based on the evidence that low levels of actinomycin D, which selectively inhibits rRNA transcription, enhances L11-HDM2 interaction, it has been proposed that L11 plays a role in the response to ribosomal perturbations to induce p53 and cell cycle arrest. In this study, we describe functional interactions of HDM2 with L23, a protein component in the 60S large ribosomal subunit. The...

Ribosomopathies: human disorders of ribosome dysfunction

“…91 MDM2 has also been shown to bind specifically to several free ribosomal proteins, including RPL5, RPL23, RPL11, RPS7, and RPL26. [92][93][94][95][96][97][98] In an elegant series of experiments, nucleolar disruption, experimentally induced by treatment with actinomycin D, was shown to lead to the release of RPL11 and other ribosomal proteins into the nucleoplasm, the binding of RPL11 to MDM2, the inhibition of MDM2 activity, and the consequent accumulation of p53. 75,95 A schematic view of this pathway is shown in Figure 3A.…”

Ribosome biogenesis surveillance: probing the ribosomal protein-Mdm2-p53 pathway

“…Nearly a decade later, in screens seeking out novel Mdm2 modulating proteins, the large subunit RPs RPL5, RPL11 and RPL23 were all reported to bind to Mdm2, block the E3 ubiquitin ligase function of Mdm2, and promote p53 accumulation (Lohrum et al, 2003;Zhang et al, 2003;Bhat et al, 2004;Jin et al, 2004). Following these initial reports, additional evidence subsequently was produced to support the roles of RPS7 (Chen et al, 2007;Zhu et al, 2009), RPL26 (Ofir-Rosenfeld et al, 2008 and RPS3 (Yadavilli et al, 2009) as Mdm2-binding partners.…”

“…In part, these observations have led to the hypothesis of the nucleolus as a central stress response regulator for p53 activation (Rubbi and Milner, 2003). One technique that has been used to induce nucleolar stress is inhibition of precursor rRNA synthesis using low doses of actinomycin D (Bhat et al, 2004;Jin et al, 2004), 5-flourouracil (Sun et al, 2007) and mycophenolic acid (Sun et al, 2008). However, with regards to an RP-Mdm2-p53 pathway, it is imperative to identify the natural biological mechanisms of nucleolar stress, particularly as they pertain to ribosome biogenesis.…”

The balance between rRNA and ribosomal protein synthesis up- and downregulates the tumour suppressor p53 in mammalian cells

“…In fact, the large subunit ribosome proteins rpL5, rpL11 and rpL23 were all reported to bind MDM2, thus inducing p53 stabilisation by inhibiting its E3 ubiquitin ligase function (Lohrum et al, 2003;Zhang et al, 2003;Bhat et al, 2004;Jin et al, 2004). The impairment of the nucleolar function has been proposed as a common denominator of many signalling pathways leading both to the suppression of MDM2 function and to p53 stabilisation and activation (Rubbi and Milner, 2003).…”