NUCLEAR HORMONE ACTION--ROLE OF UBIQUITIN PATHWAYS

类固醇、维生素、维甲酸和甲状腺激素通过它们的同源核受体对细胞的生长、发育、分化、动态平衡和肿瘤发生产生深远的影响，这些核受体构成了一个结构上相关的细胞内激素激活转录因子超家族。最近的证据表明，核激素受体(NHR)的转录调控需要一组不同的蛋白质，称为共激活因子。共激活子描述了一类不断增长的蛋白质，这些蛋白质以激素依赖的方式与受体相互作用，并且是受体最大限度地激活基因所必需的。人们普遍认为，共激活剂通过在DNA结合的受体和预起始复合体的基本转录因子之间起桥梁作用，并通过提供组蛋白乙酰转移酶活性HAT来增强受体的功能，组蛋白乙酰转移酶可以破坏局部抑制性染色质结构，有助于提高转录活性。最近，我们等克隆了泛素蛋白酶体和类泛素(NEDD8，神经前体细胞表达发育下调)途径酶、E6相关蛋白(E6-AP)、RSP5/RPF1、泛素偶联酶(UBC)、泛素激活酶(SUG1)和泛素激活酶(UBA3)作为NHR的共激活物。这些辅活化子还具有其他酶活性，如泛素活化、泛素化结合、泛素连接和蛋白酶活性，而不是HAT活性。这些观察表明，泛素和NEDD8通路在核激素受体介导的基因激活中可能起到调节作用。然而，这些途径调控核受体基因激活的确切机制目前还完全不清楚。根据我们的初步观察，我们假设泛素和NEDD8通路是核激素受体共激活的调节器。为了了解这些途径调节核激素受体基因激活的机制，我们提出了以下三个具体目的：A)鉴定和鉴定E6-AP相互作用蛋白(S)，并研究它们与E6-AP在核激素受体共激活中的作用；B)鉴定和鉴定泛素-蛋白酶体途径的靶蛋白，并了解这些途径调节核激素受体基因激活的机制。我们提出以下三个具体目的：A)鉴定和鉴定E6-AP相互作用蛋白(S)，并研究它们与E6-AP在核激素受体共激活中的作用。2)鉴定和鉴定泛素-蛋白酶体途径的靶蛋白，并了解该途径调节核激素受体基因激活的机制；3)鉴定E1泛素激活酶UBA3。研究NEDD8途径作为NHR的共同激活剂的作用，并表征这种作用在核激素受体共同激活中的作用。

Steroids, vitamins, retinoids, and thyroid hormones exert profound effects on cell growth, development, differentiation, homeostasis and tumorigenesis through their cognate nuclear receptors which make up a superfamily of structurally related intracellular hormone-activated transcription factors. Recent evidence suggests that transcriptional regulation by nuclear hormone receptors (NHRs) require a diverse group of proteins termed co-activators. Co-activators portray a growing class of proteins that interact with receptors in a hormone-dependent manner and are required for maximal gene activation by the receptors. It is the general belief that co-activators enhance receptor function by acting as a bridge between the DNA-bound receptor and basal transcription factors of the pre-initiation complex and also by providing histone acetyl transferase activity, HAT, which disrupts the local repressive chromatin structure and contributes to increase transcriptional activity. Recently, our laboratory and other have cloned ubiquitin-proteasome and ubiquitin-like (NEDD8, neural precursor cell-expressed developmentally down-regulated) pathway enzymes, E6-associated protein (E6-AP), RSP5/RPF1, ubiquitin- conjugating enzymes (UBCs), SUG1 and E1 ubiquitin-activating enzymes (UBA3) as co-activators of NHRs. These co-activators possess other enzymatic activities such as ubiquitin activation, ubiquitination conjugation, ubiquitin ligation and protease activities, instead of HAT activity. These observations suggest a possible regulatory role for the ubiquitin and NEDD8 pathways in nuclear hormone receptor-mediated gene activation. However, the exact mechanisms by which these pathways regulate nuclear receptor gene activation is completely unknown at this time. Based on our preliminary observations, we hypothesize that the ubiquitin and NEDD8 pathways are modulators of nuclear hormone receptor co-activation. In order to understand the mechanism by which these pathways modulate nuclear hormone receptor gene activation, we propose the following three specific aims: A) To identify and characterize the E6-AP interacting protein(s) and study their role along with E6-AP in nuclear hormone receptor co-activation, B) To identify and characterize target proteins of the ubiquitin-proteasome pathway and understand the mechanism by which these pathways modulate nuclear hormone receptor gene activation, we propose the following three specific aims: A) To identify and characterize the E6-AP interacting protein(s) and study their role along with E6-AP in nuclear hormone receptor co-activation, B) To identify and characterize target proteins of the ubiquitin-proteasome pathway and understand the mechanism by which this pathway modulates nuclear hormone receptor gene activation, and C) To characterize the E1 ubiquitin-activating enzyme, UBA3,. Of the NEDD8 pathway as a co-activator of NHRs and to characterize this role in nuclear hormone receptor co-activation.