C/EBP alpha in Aging Liver

描述（由申请人提供）：部分肝脏切除后，衰老的肝脏失去增殖能力，导致老年人死亡率较高。我们的实验室研究控制肝脏增殖的分子机制。肝脏特异性蛋白CCAAT/增强子结合蛋白α (C/EBPalpha)是肝脏增殖的强抑制剂。C/EBPalpha通过与细胞周期蛋白依赖性激酶2和4的直接相互作用抑制年轻肝脏的增殖。在脂肪组织中，C/EBPalpha通过抑制E2F转录导致生长停滞。我们最近发现，衰老使肝脏中的C/EBPalpha从抑制cdks转变为抑制E2F，这一途径通常在脂肪组织中起作用。在老年肝脏中，C/EBPalpha存在于高分子量复合物C/EBPalpha- rb - e2f4 - brm中。该复合物占据e2f依赖性启动子，并阻断肝脏增殖所需基因的表达激活。部分肝切除术后未能减少这种复合物似乎是老年肝脏增生反应减少的主要原因。该复合物的三个组成部分Rb、C/EBPalpha和E2F4在老年肝脏中被不同程度磷酸化。C/EBPalpha ser93位点的磷酸化是年龄特异性复合物形成的关键事件。我们最近发现cdk4和PP2A是调节C/EBPalpha的Serl93磷酸化-去磷酸化的酶。在这项应用中，我们建议研究衰老对这些信号转导途径的影响，并确定它们在C/EBPalpha-Rb-E2F4-Brm复合物的出现和增殖反应减少中的作用。我们的工作假设是：1)C/EBPalpha， E2F4和Rb的磷酸化促进了年龄相关复合物的形成；2)肝脏中C/EBPalpha Serl93的磷酸化由cdk4介导，并通过PI3K-Akt-PP2A通路的作用被逆转；3)衰老的肝脏通过减少PI3K-Akt-PP2A通路和激活cdk4来增加年龄特异性复合物。在Specific Aim 1中，我们将研究衰老激活cdk4和下调PP2A活性的机制。特异性目的2研究Rb和E2F4的磷酸化是否影响它们形成年龄特异性C/EBPalpha复合物的能力。在Specific Aim 3中，将鉴定该复合物的其他蛋白质，并研究它们在肝脏增殖中的作用。基于该项目获得的数据，我们正计划开始开发一种纠正老年人肝脏增生的策略。

DESCRIPTION (provided by applicant): An aging liver loses the ability to proliferate after partial hepatic resections leading to a higher mortality in the elderly. Our laboratory' investigates molecular mechanisms that control liver proliferation. Liver specific protein, CCAAT/Enhancer Binding Protein alpha (C/EBPalpha), is a strong inhibitor of liver proliferation. C/EBPalpha inhibits proliferation of young livers through direct interactions with cyclin dependent kinases 2 and 4. In adipose tissues, C/EBPalpha causes growth arrest via repression of E2F transcription. We have recently found that aging switches C/EBPalpha in liver from inhibition of cdks to repression of E2F, the pathway that normally operates in adipose tissues. In old livers, C/EBPalpha is observed in a high MW complex C/EBPalpha-Rb-E2F4-Brm. This complex occupies E2F-dependent promoters and blocks activation of genes whose expression is required for liver proliferation. A failure to diminish this complex after partial hepatectomy seems to be a major cause for the reduced proliferative response in old livers. Three components of the complex, Rb, C/EBPalpha and E2F4, are differentially phosphorylated in old livers. Phosphorylation of C/EBPalpha at Serl93 is a key event in the formation of the age-specific complex. We have recently identified cdk4 and PP2A as enzymes that regulate phosphorylation-dephosphorylation of Serl93 of C/EBPalpha. In this application, we propose to examine the effects of aging on these signal transduction pathways and determine their roles -in the appearance of the C/EBPalpha-Rb-E2F4-Brm complex and in the reduction of the proliferative response. Our working hypotheses are that: 1) Phosphorylation of C/EBPalpha, E2F4, and Rb promotes the formation of the age related complex, 2) Phosphorylation of Serl93 of C/EBPalpha in the liver is mediated by cdk4 and is reversed by the action of the PI3K-Akt-PP2A pathway, 3) Aging liver increases the age-specific complex by diminishing the PI3K-Akt-PP2A pathway and by activation of cdk4. In Specific Aim 1, we will examine mechanisms by which aging activates cdk4 and down-regulates activity of PP2A. Specific Aim 2 examines whether phosphorylation of Rb and E2F4 affects their ability to form the age-specific C/EBPalpha complex. In Specific Aim 3, additional proteins of the complex will be identified and their role in liver proliferation will be examined. Based on data obtained in this project, we are planning to start developing a strategy to correct liver proliferation in elderly.