Role of Uhrf1 in Liver Development, Regeneration and Carciogenesis

描述（由申请人提供）：胚胎肝脏发育和成人肝脏再生的特点是调节肝细胞增殖。这与伴随许多慢性肝病和肝细胞癌（HCC）的不受调节的肝细胞增殖形成对比。虽然有人认为，共同的遗传途径调节生理性和病理性肝细胞增殖，但只有少数哺乳动物的研究证实了这一假设。通过斑马鱼胚胎的前向遗传筛选，结合成人肝脏再生的功能研究和人类HCC样本的表达分析，我们发现含有PHD和RING finger结构域-1 （uhrf1）基因的泛素样基因对生理性肝细胞增殖至关重要。我们还认为，UHRF1表达和/或调控的改变也有助于癌症中不受控制的增殖。本提案的工作将结合斑马鱼发育和遗传学、人类癌症基因组分析、哺乳动物组织培养细胞周期研究和生物化学来解决与肝细胞增殖相关的UHRF1功能的三个方面。在特异性目标1中，我们将研究UHRF1在调节斑马鱼胚胎肝细胞增殖中的作用机制。在Specific Aim 2中，我们将阐明如何通过细胞周期蛋白依赖性激酶2的磷酸化来调节UHRF1。这种磷酸化的功能相关性将在培养细胞上使用生化和细胞生物学技术进行评估。在Specific Aim 3中，我们将确定UHRF1在肝癌发生中的作用。通过对人类HCC样本的分析，我们将评估UHRF1基因座的扩增有助于其在癌症中的上调的可能性。其次，我们将在斑马鱼中进行遗传和致癌研究，确定UHRF1对于肝脏肿瘤的形成是否必要和充分。总之，这一建议将胚胎和肝脏再生过程中控制肝细胞增殖的机制与控制肝癌发生的机制联系起来。这一建议与肝病领域有直接关系。由于肝脏疾病的负担仍然巨大，鉴定在肝细胞增殖和HCC进展中起关键作用的基因具有重要的科学和临床意义。目的是确定肝细胞增殖的新机制，以帮助开发用于治疗包括HCC在内的慢性肝病患者的化疗药物。公共卫生相关性：我们对肝脏如何发育，损伤后如何自我恢复以及肝癌如何发生感兴趣。我们相信这三个过程是相互联系的，了解正常的肝脏生长将有助于照顾肝癌患者。我们相信，我们已经发现了一种名为UHRF1的基因，它参与了这三个过程，并将研究它在每种情况下如何发挥作用。小灵通398/2590 （Rev. 09/04，重新发布4/2006）页延续格式页

DESCRIPTION (provided by applicant): Liver development in embryos and regeneration in adults are characterized by regulated hepatocyte proliferation. This contrasts with the unregulated hepatocyte proliferation that accompanies many chronic hepatic diseases and hepatocellular carcinoma (HCC). While there is suggestion that common genetic pathways regulate both physiologic and pathologic hepatocyte proliferation, only a few studies in mammals illustrate this hypothesis. By forward genetic screening in zebrafish embryos combined with functional studies on liver regeneration in adults and expression analysis of human HCC samples, we have found that the ubiquitin-like, containing PHD and RING finger domains-1 (uhrf1) gene is essential for physiologic hepatocyte proliferation. We also believe that alterations in UHRF1 expression and/or regulation also contribute to deregulated proliferation in cancer. The work in this proposal will use a combination of zebrafish development and genetics, human cancer genomic analysis and mammalian tissue culture cell cycle studies and biochemistry to address 3 aspects of UHRF1 function in relation to hepatocyte proliferation. In Specific Aim 1, we will examine the mechanism by which UHRF1 functions in regulating hepatocyte proliferation in zebrafish embryos. In Specific Aim 2, we will elucidate how UHRF1 is regulated through phosphorylation by cyclin dependent kinase 2. The functional relevance of this phosphorylation will be assessed on cultured cells using biochemical and cell biological techniques. In Specific Aim 3, we will determine the role of UHRF1 in hepatocarcinogenesis. By analysis of human HCC samples, we will evaluate the possibility that amplification of the UHRF1 locus contributes to its upregulation in cancer. Secondly, we will perform genetic and oncogenic studies in zebrafish and determine if UHRF1 is necessary and sufficient for hepatic tumor formation. In summary, this proposal will link together mechanisms that control hepatocyte proliferation in the embryo and during liver regeneration with those that control hepatocarcinogenesis. This proposal has direct relevance to the field of liver disease. Because the burden of liver disease remains enormous, the identification of genes that play critical roles in hepatocyte proliferation and in HCC progression are of significant scientific and clinical importance. The goal is to identify novel mechanisms of hepatocyte proliferation that can aid in the development of chemotherapeutic agents for use in management of patients with chronic liver diseases including HCC. PUBLIC HEALTH RELEVANCE: We are interested in how the liver develops, how it restores itself after injury and how liver cancer occurs. We believe that all three processes are linked and that understanding of normal liver growth will help in caring for patients with liver cancer. We believe that we have discovered a gene called UHRF1 that is involved in all the three processes and will study how it plays a role in each situation. PHS 398/2590 (Rev. 09/04, Reissued 4/2006) Page Continuation Format Page