HNF-4 FUNCTION IN MODELS OF TRAUMATIC INJURY

HNF-4 在创伤模型中的功能

• 批准号: 6930357
• 负责人: PETER A BURKE
• 金额: $ 35.85万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-08-01 至 2009-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6930357
• 关键词: acute phase protein; alpha 1 antitrypsin; binding sites; biological signal transduction; chromatin immunoprecipitation; gene expression; genetic regulation; immune response; immunogenetics; inflammation; laboratory mouse; laboratory rat; liver function; microarray technology; phosphorylation; protein binding; protein localization; protein purification; protein structure function; serum albumin; site directed mutagenesis; tissue /cell culture; transcription factor; trauma; two dimensional gel electrophoresis

DESCRIPTION (provided by applicant): The body's immediate reaction to serious trauma, denoted the Acute Phase Response (APR), coordinates a wide variety of inflammatory signals, and produces a common response, in the massive induction of protective proteins by the liver. While the APR is designed for survival, severe or prolonged activation with its interruption of normal homeostatic functions, likely contributes to organ failure and death in some critically ill patients. Evidence suggests that the APR's repression of steady-state liver function is a byproduct of its mode of induction. Further evidence suggests that this is a highly regulated process, which shares pathways important to early cell type development, and with some signals associated with the proliferative response. Evidence is presented that injury induced regulation of differentiated genes may be mediated through phosphorylation of liver-specific transcription factors, particularly HNF-4. Better understanding of the mechanisms regulating this process would have therapeutic importance in advantageous support of the acute phase response. The first aim will dissect where on HNF-4 phosphorylation occurs. This will be done by phosphopeptide mapping. We will also identify the kinases and thus the signal transduction pathways that are involved in initiating the APR. This will generate original materials, in the form of altered HNF-4 molecules, to test the importance of this phosphorylation to the APR. The second aim is to trace the effect of this phosphorylation on the biochemical activities of HNF-4, its DNA binding and site selection, and changes in its interactions and activities in the cell. This will be carried out using diagnostic chromatin immunoprecipitation. The third Aim will use DNA microarrays, to produce a picture of the genes regulated by HNF-4 before and after injury. An in vitro model of acute phase induction will also be used to allow direct manipulation of HNF-4 and to measure its effects on acute phase transcriptional events. These studies will elucidate the mechanisms of HNF-4 functions and test, the importance of HNF-4's modifications. It will establish what transcriptional events are common to the early phase of the APR, when normal liver function is modified, and the liver prepares for subsequent massive modulation of protein production.

描述（由申请人提供）：人体对严重创伤的直接反应，表示急性相反应（APR），协调多种炎症信号，并产生共同的反应，以大量通过肝脏诱导保护蛋白。虽然APR是为生存而设计的，但由于正常的稳态功能中断而严重或长时间激活，但可能导致某些重症患者的器官衰竭和死亡。证据表明，APR对稳态肝功能的抑制是其诱导方式的副产品。进一步的证据表明，这是一个高度调节的过程，该过程具有对早期细胞类型发育重要的途径，并且具有与增殖反应相关的一些信号。有证据表明，损伤诱导的分化基因的调节可以通过肝特异性转录因子（特别是HNF-4）的磷酸化来介导。对调节此过程的机制的更好理解将具有对急性相响应的有利支持具有治疗意义。第一个目的将剖析在HNF-4上发生磷酸化的位置。这将通过磷酸肽映射来完成。我们还将确定激酶，从而确定启动APR所涉及的信号转导途径。这将以改变的HNF-4分子的形式生成原始材料，以测试这种磷酸化对APR的重要性。第二个目的是追踪这种磷酸化对HNF-4的生化活性，其DNA结合和位点选择的影响以及其在细胞中其相互作用和活性的变化。这将使用诊断染色质免疫沉淀进行。第三个目标将使用DNA微阵列来产生受伤前后HNF-4调控的基因的图片。急性相诱导的体外模型也将用于直接操纵HNF-4，并测量其对急性相转录事件的影响。这些研究将阐明HNF-4功能和测试的机制，即HNF-4修改的重要性。当正常的肝脏功能被修饰时，它将确定哪些转录事件与APR的早期早期相同，并且肝脏为随后的大规模调节蛋白质产生做准备。