GROWTH HORMONE SIGNALING TO THE NUCLEUS

生长与复杂的遗传程序有关，涉及许多基因表达的变化。生长激素（GH）是正常生长的关键调节因子，引起基因表达的变化。在以c-fos为模型研究GH调控基因转录的机制时，我们提出GH调控c-fos中的多个DNA序列，目前已经确定了4个：与GH应答时结合信号转导和转录激活因子（Stats） 1和3的sis诱导元件（SIE），需要血清应答因子（SRF）和Elk-1介导GH促进转录的血清应答元件（SRE），一个可以结合c-fos和c- Jun的AP-1位点；CCAAT/增强子结合蛋白（C/EBP）位点，结合C/EBP β和δ。通过分析GH对与这些复合物相关的转录因子的调控，我们迄今为止已经确定了GH受体（GHR）和细胞核之间的三种不同的信号通路，由丝裂原活化蛋白激酶（MAPKs）和磷脂酰肌醇3'激酶（PI3K）介导。本研究以这些发现为出发点，探讨生长激素调控的转录涉及高阶转录因子复合物的形成和相关的染色质重塑的假设。研究人员建议研究生长激素调控基因上的转录因子如何协调成高阶复合物，以及共激活因子和共抑制因子如何参与这些复合物，从而导致染色质重塑，从而促进生长激素应答中的转录激活。本提案的第一部分描述了C/EBPbeta磷酸化状态在gh调控的C fos转录中的调节和重要性。C/EBPbeta是特别有用的，因为它与许多转录因子和共激活因子相互作用。因此，C/EBPbeta为鉴定gh调控的C -fos表达中的高阶复合物提供了一个起点。由于Stat 5参与了几种GH调控基因的调控，其中一些基因含有C/EBPbeta结合位点，因此我们将研究Stat 5是否参与GH调控的高阶转录因子复合物。这种高阶复合物的组装可能涉及协同激活剂和/或协同抑制剂分子，它们可以与生长激素调节的许多单个转录因子相互作用。在gh调节的转录复合体中，共激活因子和共抑制因子的参与以及由此产生的染色质重塑将被检查。拟议的研究将提供GH调控基因转录机制的关键信息，特定基因上响应GH的转录复合物组装的特异性，以及适用于多个基因的基因表达的协调调节。通过增加我们对正常生长调节的基本机制的理解，它们为理解异常细胞生长（如癌症）提供了基础，并为设计合理的生长控制疗法提供了基础。

Growth is associated with complex genetic programs involving changes in the expression of many genes. Growth hormone (GH), a key regulator of normal growth, elicits changes in gene expression. In examining the mechanisms by which GH regulates gene transcription using c-fos as a model, we proposed that GH regulated multiple DNA sequences in c fos, and have now identified four: the Sis-Inducible Element (SIE) which binds Signal Transducers and Activators of Transcription (Stats) 1 and 3 in response to GH, the Serum Response Element (SRE) which requires Serum Response Factor (SRF) and Elk-1 to mediate GH- promoted transcription, an AP-1 site which can bind c-Fos and c- Jun; and a CCAAT/Enhancer Binding Protein (C/EBP) site which binds C/EBPbeta and delta. By analyzing regulation by GH of the transcription factors associated with these complexes, we have so far identified three distinct signalling pathways between the GH receptor (GHR) and the nucleus, mediated by Stats by Mitogen Activated Protein Kinases (MAPKs) and by phosphatidyl inositol 3' kinase (PI3K). The present proposal takes these findings as a point of departure to investigate the hypothesis that GH- regulated transcription involves formation of higher order transcription factor complexes and related chromatin remodelling. It is proposed to examine how the transcription factors on GH- regulated genes are coordinated into higher order complexes, and how co-activators and co-repressors participate in such complexes to bring about chromatin remodelling that contributes to transcriptional activation in response to GH. The first part of this proposal delineates the regulation and importance of the phosphorylation state of C/EBPbeta in GH-regulated transcription of c fos. C/EBPbeta is particularly informative because it interacts with many transcription factors and co-activators. Thus, C/EBPbeta provides a starting point for identifying higher order complexes in GH-regulated c-fos expression. Since Stat 5 participates in the regulation of several GH-regulated genes, some of which contain C/EBPbeta binding sites, the participation of Stat 5 in higher order transcription factor complexes regulated by GH will be examined. The assembly of such higher order complexes is likely to involve co-activator and/or co- repressor molecules which can interact with many of the individual transcription factors which GH regulates. Participation of co-activators and co-repressors and the resultant chromatin remodelling in GH-regulated transcription complexes will be examined. The proposed studies will provide critical information on mechanisms for GH-regulated gene transcription, for specificity in assembly of transcription complexes in response to GH on particular genes, and on coordinated regulation of gene expression applicable to multiple genes. By adding to our understanding of fundamental mechanisms for regulation of normal growth, they provide a basis for understanding aberrant cellular growth, as in cancer, and for design of rational therapies toward growth control.