Gene Regulatory Mechanisms Controlling Tissue Maturation and Polyploidization

一、摘要 多倍体--一种细胞携带两组以上染色体的状态--在自然界中经常被观察到， 然而，控制倍性的遗传机制及其功能意义仍然是个谜。肝脏，在 特别是在出生后发育期间获得高比例的多倍体肝细胞；以及 肝脏多倍化的频率和程度在损伤、DNA损伤和 氧化应激，但在肝细胞癌中降低。最近的证据表明，多倍体 通过减缓肝细胞的增殖能力和维持一种 肿瘤抑制因子的储存库。然而，人们对支配分子事件的了解很少。 肝脏多倍化的出生后启动/促进或染色体倍性差异如何影响 肝细胞的转录和转录后活性。我们之前已经证明了 RNA结合蛋白ESRP2是一个关键的发育调节因子，它激活了成人剪接 促进肝细胞终末分化、功能能力和成熟的计划。目标 这项建议的目的是(I)确定ESRP2及其剪接的生理必要性/充分性- 调控网络在驱动肝细胞多倍体中的作用，以及(Ii)定义了 肝细胞转录产量的倍性。AIM 1将使用ESRP2获得和丧失功能的小鼠模型来 确定通过ESRP2激活的RNA剪接中的程序性变化是否对 肝细胞多倍化。在目标2中，我们将从二倍体和 多倍体小鼠肝细胞研究倍性如何影响稳态水平和替代 全基因组水平上肝脏转录的剪接模式。提议的目标将检测新基因。 控制多倍化的监管机制(S)，同时发现以前未被认识到的 选择性剪接和细胞多倍体。

I. ABSTRACT Polyploidy—a state in which cells carry more than two sets of chromosomes—is frequently observed in nature, yet, the genetic mechanisms controlling ploidy and its functional significance remain enigmatic. The liver, in particular, gains a high percentage of polyploid hepatocytes during postnatal period of development; and the frequency and extent of hepatic polyploidization are further increased following injury, DNA damage, and oxidative stress, but are decreased in hepatocellular carcinoma. Recent evidence suggests that polyploidy safeguards the liver from tumorigenesis by slowing the proliferative capacity of hepatocytes and maintaining a reservoir of tumor suppressors. However, there is minimal understanding of the molecular events that govern the postnatal initiation/promotion of hepatic polyploidization or how differences in chromosomal ploidy affect the transcriptional and posttranscriptional activities of hepatocytes. We have previously demonstrated that the RNA binding protein ESRP2 is a key developmentally regulated factor, which activates an adult splicing program to facilitate terminal differentiation, functional competence, and maturation of hepatocytes. The goals of this proposal are to (i) determine the physiological necessity/sufficiency of ESRP2 and its splicing- regulatory-network in driving hepatocyte polyploidy, and (ii) define the quantitative and qualitative impact of ploidy on hepatocyte transcriptional output. Aim 1 will use ESRP2 gain-and loss-of-function mouse models to determine if programmed changes in RNA splicing through ESRP2 activation are crucial for the polyploidization of hepatocytes. In Aim 2, we will generate high-resolution transcriptomes from diploid and polyploid murine hepatocytes to investigate how ploidy influences the steady-state levels and alternative splicing patterns of hepatic transcripts at a genome-wide scale. The proposed aims will examine new gene regulatory mechanism(s) controlling polyploidization while uncovering previously unrecognized links between alternative splicing and cellular polyploidy.