Amino Acid Regulation of the Fos/Jun Transcription Factors

描述(申请人提供)：蛋白质剥夺不仅对基因表达有直接的调节作用，还可以通过染色质相关的机制产生长期的影响。然而，关于饮食蛋白质波动对包括转录在内的基本细胞功能的影响，我们的认识存在显著差距。在单个组织/细胞水平上，蛋白质缺乏反映为氨基酸的限制，因此，培养细胞的氨基酸缺失是研究转录调控基因组机制的有用模型。体内蛋白质限制或细胞培养中AA缺乏可激活多途径AA反应(AAR)，这是最终导致ATF4合成的几种应激激活信号机制之一。一项微阵列研究发现，FOS/JUN转录因子家族的成员是人肝癌细胞中AA调节的基因。随后的研究表明，cJUN调控着几个已知的AAR靶点的下游基因。还提出了另外两个新的观察结果。首先，cJUN基因的诱导不依赖于ATF4，因此必须包含一个新的AA反应元件(AARE)，该元件可能与新的信号通路有关。其次，cJUN的诱导发生在人肝癌细胞中，而不是在原代人肝细胞或未转化的永生化肝细胞系中。提出的研究具有创新性，因为：1)他们将通过ATF4不依赖的机制首次对蛋白质/AA反应基因进行研究，2)他们将在体内的动物肿瘤模型以及培养的人肝癌细胞和正常肝细胞中建立cJUN基因的调控机制。我们的全球假设是，cJUN基因包含一个新的AARE，并且在肝癌细胞中通过ATF4非依赖的信号通路被激活，并且在饮食蛋白或AA限制的过程中，cJUN的高表达支持肝癌的持续增殖。拟议的实验将检验以下假设：(假设I)cJUN基因是一个新的ATF4非依赖AA信号通路的靶标；(假设II)cJUN基因显示出新的AA反应转录机制；(假设III)cJUN在蛋白质/AA限制期间调节AAR基因和细胞增殖。 公共卫生相关性：充足的膳食蛋白质/氨基酸供应是维持健康的重要因素，特别是在怀孕期间、胎儿/新生儿发育期间，以及包括糖尿病和癌症在内的各种疾病的发展过程中。尽管有这些重要的关系，但饮食中蛋白质/氨基酸的波动对基本细胞过程和病理状态的影响，如肿瘤的发生和维持，我们知之甚少。这项拟议的研究将调查导致FOS/Jun蛋白家族的机制，这些蛋白是基因表达和细胞增殖的主要调节因子，由人类肿瘤细胞的氨基酸限制诱导，而不是正常的肿瘤细胞。

DESCRIPTION (provided by applicant): Protein deprivation not only has immediate regulatory effects on gene expression, but can also have long-term effects through chromatin-associated mechanisms. However, there are significant gaps in our knowledge about the impact of dietary protein fluctuation on basic cell functions, including transcription. At the level of individual tissues/cells, dietary protein deficiency is reflected as amino acid (AA) limitation and therefore, AA deprivation of cultured cells is a useful model to investigate the genomic mechanisms of transcriptional control. Dietary protein limitation in vivo or AA deprivation in cell culture activates a multi-pathway AA response (AAR) that is one of several stress-activated signaling mechanisms that culminate in ATF4 synthesis. A microarray study identified members of the FOS/JUN family of transcription factors as AA-regulated genes in human HepG2 hepatocellular carcinoma (HCC) cells. Subsequent studies showed that cJUN regulates several downstream genes that are known AAR targets. Two additional novel observations were made. First, the induction of the cJUN gene is ATF4-independent, and therefore, must contain a novel AA response element (AARE), which may be linked to a novel signaling pathway. Second, the induction of cJUN occurs in human HCC cells, but not in primary human hepatocytes or in a non-transformed immortalized hepatocyte cell line. The proposed studies are innovative because: 1) they will provide the first investigation of a protein/AA responsive gene by an ATF4-independent mechanism, and 2) they will establish the mechanism for regulation of the cJUN gene in an animal tumor model in vivo, as well as in cultured human hepatoma cells and normal hepatocytes. Our global hypothesis is that the cJUN gene contains a novel AARE and is activated in HCC cells by an ATF4-independent signaling pathway, and that elevated cJUN expression supports continued HCC proliferation during dietary protein or AA limitation. The proposed experiments will test the following hypotheses: (Hypothesis I) The cJUN gene is the target of a novel ATF4-independent AA signaling pathway; (Hypothesis II) The cJUN gene exhibits novel AA-responsive transcriptional mechanisms; and (Hypothesis III) cJUN modulates AAR genes and cell proliferation during protein/AA limitation. PUBLIC HEALTH RELEVANCE: Sufficient dietary protein/amino acid availability is an important factor in maintenance of health, especially during pregnancy, fetal/neonatal development, and in the progression of a wide range of diseases, including diabetes and cancer. Despite these important relationships, the influence of dietary protein/amino acid fluctuation on fundamental cellular processes and pathological states, such as the development and maintenance of tumors, are topics about which we know relatively little. The proposed research will investigate the mechanisms that lead the FOS/JUN family of proteins, major regulators of gene expression and cell proliferation, to be induced by amino acid limitation of human tumor cells, but not their normal counterparts.