Nuclear receptors: action, functions, and roles in disease

1.RORpha和Gamma：维甲酸相关的孤儿受体a和g(RORA和RORg)是核受体超家族的成员。为了确定RORA和g的生理功能，对RORA和g功能缺陷的小鼠进行了分析。RORg在免疫系统中发挥多种功能。RORg的表达是淋巴器官发生所不可缺少的，对胸腺细胞的动态平衡起着关键作用。最近发现了RORg在Th17细胞分化中的作用。我们证明了RORA和RORg都是在Th17细胞分化过程中诱导的，并且双基因敲除小鼠对实验性自身免疫性脑脊髓炎具有抵抗力，这是多发性硬化症的一个模型。此外，RORg缺陷小鼠不太容易受到OVA(OVA)诱导的小鼠炎症的影响，OVA是过敏性呼吸道疾病的模型。 维甲酸相关孤儿受体α(RORA)和伽马受体(RORg)在肝脏中均有表达，但其在肝脏中的生理功能尚未明确。RORA1和RORg1在昼夜节律中表现出振荡的表达模式。通过基因芯片分析比较WT、RORA缺陷交错鼠(RORasg/sg)、RORg-/-和RORasg/sgRORg-/-双基因敲除(DKO)小鼠肝脏的基因表达谱表明，RORA和RORg在编码几种I相和II相代谢酶的基因调控中特别重要，包括几种3b-羟基类固醇脱氢酶(Hsd3b)、细胞色素P450(Cyp)酶和磺基转移酶。此外，我们的结果表明，RORA和RORg各自影响一组特定基因的表达，但也表现出功能冗余。我们的研究表明，RORA和RORg受体影响几种代谢途径的调节，包括参与类固醇、胆汁酸和外源生物代谢的途径，这表明ROR在控制代谢动态平衡方面起着重要作用。RORA在能量稳态的调节中起着重要作用。RORA基因缺陷的小鼠对饮食诱导的肥胖和减少脂肪组织的炎症具有抵抗力。已鉴定出一些受RORA正调控的致脂基因。由于RORs作为配体依赖的转录因子发挥作用，RORA可能成为肥胖管理的治疗靶点。研究发现，RORg在生物钟下游的几个基因的昼夜节律调节中起着重要作用。 2.TAK1：核孤儿受体TAK1既是转录的正性调节因子，也是负性转录调节因子，但对影响其活性的因素知之甚少。酵母双杂交分析以TAK1的配体结合域为诱饵，鉴定出一种新的TAK1相互作用蛋白，称为TIP27。我们的研究表明，TIP27是TAK1转录激活的有效抑制因子，因此可能在TAK1对多种生理功能的调节中发挥关键作用。一代TAK1基因敲除小鼠揭示了目前正在研究的几种表型。TAK1缺陷(TAK1-/-)小鼠，并报告这些小鼠表现出较小的小脑和VI-VII小叶的叶化缺陷。TAK1的缺失导致小脑颗粒神经元、浦肯野细胞和放射状胶质细胞在发育过程中的协调缺陷，改变了长期的神经行为功能。这些数据表明，TAK1是神经发育调节行为的重要转录调节因子。此外，我们还发现TAK1在能量动态平衡的调节中起作用。TAK1基因缺陷的小鼠可以预防肥胖、肝脏脂肪变性和胰岛素抵抗。已鉴定出一些受TAK1正调控的致脂基因。 受体相关蛋白(RAP80)是一种含有两个泛素相互作用基序(UIM)的核蛋白，它与雌激素受体α(ERA)以激动剂依赖的方式相互作用。此外，RAP80与DNA修复有关，与肿瘤抑制因子乳腺癌-1(BRCA1)蛋白复合体相关，并介导BRCA1易位到DNA损伤部位。我们发现这种易位依赖于RAP80的UIM。我们证明了共济失调-毛细血管扩张突变蛋白激酶(ATM)在体外能够在Ser205处磷酸化RAP80。利用RAP80Ser205P抗体特异性识别RAP80在Ser205处的磷酸化，我们证明了RAP80Ser205P易位到DNA损伤的位置。我们证明这种磷酸化是由ATM介导的，不需要功能上的BRCA1。紫外线(UV)照射也能诱导RAP80移位到与H_2AX共定位的DNA损伤部位。我们进一步证明，这种易位也依赖于RAP80的UIM，并且紫外线诱导的RAP80在Ser205的磷酸化是由ATR而不是ATM介导的。我们的发现表明，RAP80在不同类型的DNA损伤反应信号通路中具有更普遍的作用。利用基因组和功能分析，我们证实了RAP80基因的表达是由主调控因子P53以DNA损伤反应的方式调节的。这种调节是通过RAP80启动子中的一个非规范的P53反应元件在转录水平上进行的。虽然RAP80可以被P53诱导，但RAP80也能够通过与P53和E3泛素连接酶Hdm2的结合来调节P53，提供Hdm2依赖的P53多泛素化增强。因此，我们提供的证据表明，RAP80可以在由RAP80、Hdm2和P53主调控网络组成的自动调节环中发挥作用，这意味着这个环在基因组稳定和肿瘤发生中发挥重要作用。

I. RORalpha and gamma: The retinoid-related orphan receptor a and g (RORa and RORg) are members of the nuclear receptor superfamily. To identify the physiological functions of RORa and g, mice deficient in RORa and g function were analyzed. RORg exhibit several functions in the immune system. RORg expression is indispensable for lymph node organogenesis and plays a critical role in thymocyte homeostasis. Recently a role for RORg in Th17 cell differentiation was identified. We demonstrated that both RORa and RORg are induced during Th17 cells differentiation and double knockouts mice are resistant to experimental autoimmune encephalomyelitis, a model for multiple sclerosis. In addition, RORg-deficient mice are less susceptible to aovalbumin (OVA)-induced inflammation in mice, a model for allergic airway disease. Retinoid-related orphan receptors alpha (RORa) and gamma (RORg) are both expressed in liver; however, their physiological functions in this tissue have not yet been clearly defined. RORa1 and RORg1 show an oscillatory pattern of expression during circadian rhythm. Comparison of gene expression profiles of livers from WT, RORa-deficient staggerer mice (RORasg/sg), RORg-/-, and RORasg/sgRORg-/- double knockout (DKO) mice by microarray analysis demonstrated that RORa and RORg are particularly important in the regulation of genes encoding several Phase I and Phase II metabolic enzymes, including several 3b-hydroxysteroid dehydrogenases (Hsd3b), cytochrome P450 (Cyp) enzymes, and sulfotransferases. In addition, our results indicate that RORa and RORg each affect the expression of a specific set of genes but also exhibit functional redundancy. Our study shows that RORa and RORg receptors influence the regulation of several metabolic pathways, including those involved in the metabolism of steroids, bile acids, and xenobiotics, suggesting that RORs are important in the control of metabolic homeostasis. RORa plays an important role in the regulation of energy homeostasis. Mice deficient in RORa are resistant to diet-induced obesity and reduced inflammation in adipose tissue. A number of lipogenic genes were identified that are positively regulated by RORa. Because RORs function as ligand-dependent transcription factors, RORa might be a therapeutic target for the management of obesity. RORg was found to play an important role in the circadian regulation of several genes downstream of the circadian clock. II. TAK1: The nuclear orphan receptor TAK1 functions as a positive as well as a negative regulator of transcription; however little is know about factors mediating its activity. Yeast two-hybrid analysis using the ligand binding domain of TAK1 as bait identified a novel TAK1-interacting protein, referred to as TIP27. Our studies indicate that TIP27 is an effective repressor of transcriptional activation by TAK1 and, therefore, may play a critical role in the regulation of several physiological functions by TAK1. Generation of TAK1 knockout mice revealed several phenotypes that are currently being investigated. TAK1-deficient (TAK1-/-) mice and report that these mice exhibit a smaller cerebellum and deficit in foliation of lobules VI-VII. The absence of TAK1 results in a coordinated deficit in cerebellar granule neurons, Purkinje cells, and radial glia during development altering long-term neurobehavioral functioning. These data indicate that TAK1 is an important transcriptional modulator of neurodevelopmentally-regulated behavior. In addition, we found that TAK1 plays a role in the regulation of energy homeostasis. Mice deficient in TAK1 are protected against the development of obesity,hepatic steatosis,and insulin resistance. A number of lipogenic genes were identified that are positively regulated by TAK1. III. Receptor associated protein (RAP80), a nuclear protein containing two ubiquitin-interacting motifs (UIMs), interacts with the esstrogen receptor alpha (ERa) in an agonist dependent manner. In addition, RAP80 is implicated in DNA repair and is associated with the tumor suppressor Breast cancer-1 (BRCA1) protein complex and mediates BRCA1 translocation to sites of DNA damage. We showed that this translocation is dependent on the UIMs of RAP80. We demonstrated that the ataxia-telangiectasia mutated protein kinase (ATM) can phosphorylate RAP80 in vitro at Ser205. Using an anti-RAP80Ser205P antibody that specifically recognizes RAP80 phosphorylated at Ser205 we demonstrated that RAP80Ser205P translocates to sites of DNA damage. We show that this phosphorylation is mediated by ATM and does not require a functional BRCA1. Ultraviolet (UV) irradiation also induces translocation of RAP80 to DNA damage foci that co-localize with γ-H2AX. We further show that this translocation is also dependent on the UIMs of RAP80 and that the UV-induced phosphorylation of RAP80 at Ser205 is mediated by ATR, not ATM. Our findings suggest that RAP80 has a more general role in different types of DNA damage response signaling pathways. Using genomic and functional analysis we established that the expression of the RAP80 gene is regulated in a DNA damage-responsive manner by the master regulator p53. This regulation occurs at the transcriptional level through a noncanonical p53 response element in the RAP80 promoter. While it is inducible by p53, RAP80 is also able to regulate p53 through an association with both p53 and the E3 ubiquitin ligase HDM2, providing HDM2-dependent enhancement of p53 polyubiquitination. Thus, we provide evidence that RAP80 can function in an autoregulatory loop consisting of RAP80, HDM2 and the p53 master regulatory network, implying an important role for this loop in genome stability and oncogenesis.