Dale S. Haines

A Novel Cellular Protein (MTBP) Binds to MDM2 and Induces a G1 Arrest That Is Suppressed by MDM2

The MDM2 protein, through its interaction with p53, plays an important role in the regulation of the G1 checkpoint of the cell cycle. In addition to binding to and inhibiting the transcriptional activation function of the p53 protein, MDM2 binds, inter alia, to RB and the E2F-1·DP-1 complex and in so doing may promote progression of cells into S phase. Mice transgenic for Mdm2 possess cells that have cell cycle regulation defects and develop an altered tumor profile independent of their p53 status. MDM2 also blocks the growth inhibitory effects of transforming growth factor-β1 in a p53-independent manner. We show here that a novel growth regulatory molecule is also the target of MDM2-mediated inhibition. Using a yeast two-hybrid screen, we have identified a gene that encodes a novel cellular protein (MTBP) that binds to MDM2. MTBP can induce G1 arrest, which in turn can be blocked by MDM2. Our results suggest the existence of another growth control pathway that may be regulated, at least in part, by MDM2.

Analysis of p53 and mdm-2 expression in 18 patients with Sézary syndrome.

Sézary syndrome is a leukaemic form of cutaneous T‐cell lymphoma which presents with multiple cytogenetic abnormalities and responds poorly to chemotherapy. Because of the importance of the p53 tumour suppressor in maintaining genomic stability and in sensitizing transformed cells to DNA damaging agents, we looked for alterations which may affect p53 functions in 18 patients with Sézary syndrome. Cytogenetic analysis suggested frequent p53 gene inactivation since 6/18 patients had loss of one copy of 17p. However, single‐strand conformational polymorphism (SSCP) revealed that p53 gene mutations are relatively rare, occurring in only two of 18 Sézary patients. Neither of these two patients was missing a copy of 17p. Possible abnormalities of p53 pathway function through mdm‐2 over‐expression were also investigated. Although all 18 patients had normal levels of mdm‐2 RNA, 4/18 over‐expressed mdm‐2 protein. One patient with advanced disease and the highest percentage of malignant cells overexpressed mdm‐2 protein and possessed a nonsense p53 gene mutation. The five patients with abnormalities of p53 or mdm‐2 were found to have significantly higher absolute lymphocyte counts and higher absolute numbers of Sézary cells (P = 0.021 and 0.027 respectively). In summary, molecular alterations of 17p and potential p53 pathway abnormalities are a common event in Sézary syndrome and appear to be associated with more advanced disease.

B55α PP2A holoenzymes modulate the phosphorylation status of the RB-related protein p107 and its activation

Pocket proteins negatively regulate transcription of E2F-dependent genes and progression through the G0/G1 transition and the cell cycle restriction point in G1. Pocket protein repressor activities are inactivated via phosphorylation at multiple Pro-directed Ser/Thr sites by the coordinated action of G1 and G1/S cyclin-dependent kinases. These phosphorylations are reversed by the action of two families of Ser/Thr phosphatases: PP1, which has been implicated in abrupt dephosphorylation of retinoblastoma protein (pRB) in mitosis, and PP2A, which plays a role in an equilibrium that counteracts cyclin-dependent kinase (CDK) action throughout the cell cycle. However, the identity of the trimeric PP2A holoenzyme(s) functioning in this process is unknown. Here we report the identification of a PP2A trimeric holoenzyme containing B55α, which plays a major role in restricting the phosphorylation state of p107 and inducing its activation in human cells. Our data also suggest targeted selectivity in the interaction of pocket proteins with distinct PP2A holoenzymes, which is likely necessary for simultaneous pocket protein activation.