MDM2 and MDMX function together as the p53 E3 ligase

描述(申请人提供)：作为细胞应激反应的主要调节因子，P53在细胞命运决定中起着重要作用。因此，必须对P53的活性进行精确调控。在参与p53调控的众多蛋白中，MDM2和MDMX是关键的参与者，事实证明，在小鼠中，MDM2或MDMX的敲除导致了P53依赖的致死。然而，对于为什么在p53调控中同时需要MDM2和MDMX，以及MDM2和MDMX介导的p53调控的分子机制仍不完全清楚，人们知之甚少。其他人和我们已经证明，MDM2和MDMX形成一个异质复合体，更重要的是，它们在p53调控中相互依赖。结构和生化证据进一步表明，MDM2/MDMX异质复合体的形成是有利的。我们的假设是，异源复合体代表了p53控制中MDM2和MDMX的生理形式。拟议的研究将使用小鼠模型来直接检验这一假设。我们还将充分描述MDM2/MDMX异源复合体在p53泛素化和调控p53应激反应中的作用。其具体目的是：1)利用动物模型研究MDM2/MDMX复合体在p53调控中的作用；2)表征MDM2/MDMX复合体介导的P53泛素化；3)研究调控MDM2/MDMX复合体的细胞机制。随着基因敲入小鼠的成功产生，我们处于非常有利的地位，可以开展拟议的研究。这项拟议工作的发现不仅有望阐明MDM2和MDMX的非冗余功能，而且还将提供一种新的p53调控机制。 公共卫生相关性：这项应用旨在通过动物模型直接测试MDM2/MDMX异源复合体在p53控制中的重要性。同时，MDM2/MDMX复合体介导的p53泛素化将得到充分的表征，并将研究MDM2/MDMX调控的细胞机制。通过这些研究，我们将检验MDM2/MDMX异源复合体是P53的生理性E3连接酶的假设，如果被证实，这不仅将代表一种新的P53调控范式，还将阐明MDM2和MDMX的非冗余功能。此外，MDM2/MDMX复合体在p53调控中的关键作用为P53的激活提供了一个理想的靶点，例如异源复合体的解离，这可能具有治疗意义。

DESCRIPTION (provided by applicant): As a master regulator of cellular stress response, p53 plays an important role in cell fate decisions. The activity of p53 therefore must be precisely regulated. Among numerous proteins involved in p53 control, MDM2 and MDMX are the key players, as evidenced by the fact that knockout of either mdm2 or mdmx in mice resulted in p53-dependent lethality. However, little is known about why both MDM2 and MDMX are required in p53 control, and the molecular mechanisms underlying MDM2 and MDMX-mediated p53 regulation remain not fully defined. Others and we have shown that MDM2 and MDMX form a heterocomplex and, more importantly, that they depend on each other in p53 regulation. Structural and biochemical evidence further indicate that formation of the MDM2/MDMX heterocomplex is favored. Our hypothesis is that the heterocomplex represents the physiological form of MDM2 and MDMX in p53 control. The proposed studies will use mouse models to directly test this hypothesis. We will also fully characterize the MDM2/MDMX heterocomplex in p53 ubiquitination and in regulation of the p53 response to stress. The specific aims are: 1) using animal models to examine a role of the MDM2/MDMX complex in p53 regulation; 2) characterize the MDM2/MDMX complex-mediated p53 ubiquitination; 3) investigate cellular mechanisms that regulate the MDM2/MDMX complex. With the knock-in mice being successfully generated, we are very favorably positioned to carry out the proposed studies. The findings obtained from the proposed work are expected not only to shed light on the non-redundant function of MDM2 and MDMX, but also to provide a novel mechanism of p53 regulation. PUBLIC HEALTH RELEVANCE: This application aims to directly test the importance of the MDM2/MDMX heterocomplex in p53 control by using animal models. In parallel, the MDM2/MDMX complex-mediated p53 ubiquitination will be fully characterized and cellular mechanisms of MDM2/MDMX regulation will be investigated. Through these lines of investigation, we will test the hypothesis that the MDM2/MDMX heterocomplex is the physiological E3 ligase for p53, which, if proven, will not only represent a new paradigm of p53 regulation but also shed lights on the non-redundant function of MDM2 and MDMX. In addition, the pivotal role of the MDM2/MDMX complex in p53 control presents an ideal target for p53 activation, e.g. disassociation of the heterocomplex, which may have therapeutic implications.