PAQR3在结肠直肠癌发生过程中的功能以及对PI3K的空间调控研究
文献类型：学位论文
| 作者 | 王笑 |
| 学位类别 | 博士 |
| 答辩日期 | 2012-11 |
| 授予单位 | 中国科学院上海生命科学研究院营养科学研究所 |
| 授予地点 | 中国科学院上海生命科学研究院 |
| 导师 | 陈雁 |
| 关键词 | 孕酮和脂联素受体3 结肠直肠癌 结肠腺瘤样息肉基因 胰岛素敏感性 磷酸酰肌醇－3激酶 |
| 其他题名 | The Functions of PAQR3 in the Tumorigenesis of Colorectal Cancers and Spatial Regulation of PI3K |
| 学位专业 | 生物化学与分子生物学 |
| 中文摘要 | PAQR3是孕酮和脂联素受体基因家族中的一员。研究表明，PAQR3是一个定位在高尔基体上的蛋白，它可以空间调控Raf激酶，从而抑制Ras/Raf/MEK/ERK信号通路。然而，目前没有任何PAQR3在结肠直肠癌发生发展过程中作用的研究。ApcMin/+小鼠有结肠腺瘤样息肉基因（adenomatous polyposis coli，APC）的突变，会在小肠和大肠中自发形成腺瘤。为了研究Paqr3基因在结肠直肠癌发生中的作用，我们将Paqr3基因敲除小鼠和ApcMin/+小鼠杂交，发现将Paqr3敲除后，ApcMin/+小鼠的生存时间缩短，小肠中腺瘤数目和负荷增加。在结肠癌细胞系SW-480中，过表达PAQR3抑制细胞的增殖和锚定非依赖生长能力，降低表皮生长因子（Epidermal Growth Factor，EGF）诱导的细胞外信号相关激酶（Extracellular signal Related Kinase，ERK）的活化以及β-连锁蛋白（β-catenin）的入核。相反，敲减内源PAQR3促进SW-480细胞增殖和克隆形成，增加表皮生长因子刺激的细胞外信号相关激酶的磷酸化以及β-连锁蛋白在细胞核内的聚积。在人结肠直肠癌肿瘤样本中，PAQR3在癌组织中表达量显著性的低于癌旁正常组织。另外，在男性病人样本中，PAQR3的表达量和肿瘤的分级呈负相关。这些结果，首次揭示了PAQR3具有抑制结肠直肠癌的发生发展的功能。 2型糖尿病一个最重要的生理特征是胰岛素抵抗，即外周组织胰岛素信号通路和功能的降低。磷脂酰肌醇3激酶（Phosphoinositide 3-kinase ，PI3K）在胰岛素信号转导过程中起到了承上启下的作用。p110?是IA型PI3K家族的催化亚基，是已知的胰岛素信号通路中最重要的PI3K成员。我们发现了PAQR3可以空间调控p110?亚基。PAQR3和脂联素受体属于同一个家族，但是与其不同的是，PAQR3特异性定位在高尔基体上。Paqr3-/-小鼠抵抗高脂和瘦素基因缺失导致的肥胖、胰岛素抵抗以及脂肪肝。在肝原代细胞，肝组织和骨骼肌中，敲除PAQR3增加胰岛素刺激的AKT和GSK3?的磷酸化，但是对IR?和IRS-1的磷酸化没有影响。PAQR3和p110?相互作用，过表达PAQR3增加p110?在高尔基体上的定位，而敲减PAQR3降低p110?在高尔基体上的分布。PAQR3与p110?的p85结合域相互作用，过表达PAQR3剂量依赖性地降低PI3K调节亚基p85?和p110?的结合。在肝细胞中，敲除PAQR3增加胰岛素刺激的PIP3的产生和PI3K的酶活；过表达PAQR3降低胰岛素诱导的PIP3的产生和PI3K的酶活。因此，PAQR3通过和p110?相互作用，阻碍其与p85?的结合，从而起到抑制胰岛素信号通路的作用。 |
| 索取号 | D2012-142 |
| 英文摘要 | PAQR3 is a member of the progestin and adipoQ receptor (PAQR) family and was recently characterized as a spatial regulator that negatively modulates Ras/Raf/MEK/ERK signaling cascade. However, little is known about the physiological functions of PAQR3 in the tumorigenesis of colorectal cancers. The function of PAQR3 in colorectal cancer development in mice was analyzed by crossing Paqr3-depleted mice with ApcMin/+ mice that have a germline mutation of the gene-encoding tumor suppressor adenomatous polyposis coli (APC). The survival time and tumor area in the small intestine of the ApcMin/+ mice was significantly aggravated by Paqr3 deletion. The cell proliferation rate, anchorage-independent growth, EGF-stimulated ERK phosphorylation and EGF-induced nuclear accumulation of β-catenin were inhibited by PAQR3 overexpression and enhanced by PAQR3 knockdown in SW-480 colorectal cancer cells. In humans, the expression level of PAQR3 was significantly decreased in colorectal cancer samples in comparison with adjacent normal tissues. In addition, the expression level of PAQR3 was inversely associated with tumor grade in the colorectal cancer samples. Collectively, our data reveal for the first time that PAQR3 has a tumor suppressor activity in the development of colorectal cancers. One of the fundamental pathophysiological features of type 2 diabetes is insulin resistance that is characterized as reduced insulin signaling and action in peripheral tissues. Phosphoinositide 3-kinase (PI3K) plays a central role in mediating insulin signaling cascade by relaying signals from insulin receptors to downstream targets such as AKT. Meanwhile, p110????one of the catalytic subunits of class IA PI3K family, has been found to be the primary insulin-responsive PI3K implicated in insulin signaling. We demonstrate here a new mode of spatial regulation of PI3K p110? subunit by PAQR3, a close homologue of adiponectin receptors. Different from adiponectin receptors, PAQR3 is exclusively localized at the Golgi apparatus. Mice deleted of Paqr3 are resistant to obesity and insulin resistance induced by either high fat diet (HFD) or leptin deficiency in ob/ob mice. Deletion of PAQR3 enhances insulin-stimulated phosphorylation of AKT and glycogen synthase kinase 3?, but not phosphorylation of IR and IR substrate-1 (IRS-1), in hepatocytes, mouse liver, and skeletal muscle. PAQR3 is able to interact with p110?. Intracellular localization of p110? at the Golgi apparatus is increased by PAQR3 overexpression and reduced by PAQR3 downregulation. Furthermore, PAQR3 interacts with the domain of p110? involved in its binding with p85, the regulatory subunit of PI3K. Overexpression of PAQR3 dose-dependently reduces the interaction of p85? with p110?. Insulin-stimulated PI3K activity and phosphoinositide (3-5)-tri-phosphate production are enhanced by deletion and reduced by PAQR3 overexpression in hepatocytes. Thus, PAQR3 negatively regulates insulin signaling by shunting cytosolic p110? to the Golgi apparatus while competing with p85 subunit in forming a PI3K complex with p110?. |
| 语种 | 中文 |
| 源URL | [http://202.127.25.144/handle/331004/111]  |
| 专题 | 中国科学院上海生命科学研究院营养科学研究所_信号转导与营养相关疾病研究组 |
| 推荐引用方式 GB/T 7714 | 王笑. PAQR3在结肠直肠癌发生过程中的功能以及对PI3K的空间调控研究[D]. 中国科学院上海生命科学研究院. 中国科学院上海生命科学研究院营养科学研究所. 2012. |
入库方式： OAI收割
来源：上海营养与健康研究所
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