Analysis of beclin 1 in autophagy and tumor suppression

beclin 1在自噬和肿瘤抑制中的分析

基本信息

批准号：
7911042
负责人：
BETH C LEVINE
金额：
$ 13.78万
依托单位：
UT SOUTHWESTERN MEDICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-08-01 至 2010-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7911042
关键词：
1-Phosphatidylinositol 3-Kinase Abnormal Cell Address Autophagocytosis Award B Cell Proliferation Binding Biological Process Breast Carcinoma Caenorhabditis elegans Cell model Complex Cytoplasmic Organelle Development Distal Dominant-Negative Mutation Embryo Epithelial Funding Gene Deletion Gene Silencing Generations Genes Grant Growth Human In Vitro Investigation Knock-in Mouse Laboratories Life MCF7 cell Malignant Epithelial Cell Mammalian Cell Mammary gland Mediating Molecular Mus Mutant Strains Mice Nematoda Oncogenic Pathway interactions Phenotype Phosphoric Monoester Hydrolases Process Proteins RNA Interference Role Staging Structure Testing Tumor Suppression Tumor Suppressor Genes Tumor Suppressor Proteins Tumorigenicity Vesicle Xenograft Model Yeasts cell growth embryonic stem cell in vivo loss of function mutation mutant myotubularin overexpression stem tumorigenesis

项目摘要

DESCRIPTION (provided by applicant): The lysosomal pathway of autophagy is the major regulated catabolic mechanism for degrading long-lived cellular proteins and cytoplasmic organelles. Yet, until recently, very little was known about the molecular mechanisms or biological functions of autophagy. Under funding from this award, our laboratory identified the first mammalian autophagy gene, beclin 1 (appendix item #1), and demonstrated that it is a haploinsufficient tumor suppressor gene (appendix item #2). We have also found that beclin 1 and other autophagy genes are essential for a developmental arrest phenotype in C. elegans that is negatively regulated by the nematode equivalent of the mammalian oncogenic Class I PI3 kinase pathway (appendix item #3). Together, these findings raise the strong possibility that autophagy functions as a tumor suppressor mechanism. In this renewal application, we will evaluate this concept in the following two specific aims. In the first aim, we will test the hypothesis that beclin 1 exerts tumor suppressor activity through a Class III PI3 kinase-dependent mechanism that involves the induction of autophagy. To accomplish this aim, we will use a previously established human MCF7 breast carcinoma cell model to determine whether the Beclin 1-binding partner, Class III PI3-K/hVps34, (involved in autophagic vesicle nucleation) and whether autophagy genes involved in a later stage of autophagy (autophagic vesicle expansion and completion) are required for the tumor suppressor function of beclin 1. We will also test the importance of Beclin 1-Vps34 binding in tumorigenesis in vivo by studying tumorigenesis in a knock-in mouse that contains a Vps34-binding defective mutant of Beclin 1. In the second aim, we will test the hypothesis that autophagy genes downstream of beclin 1 are required for negative growth control and tumor suppression. To accomplish this aim, we will study (1) spontaneous tumorigenesis in mice with heterozygous deletion of atg3, atg5, and atg7; (2) the in vivo tumorigenicity of embryonic stem (ES) cells with homozygous deletions of the autophagy genes, atg3, atg5 and beclin 1, and (3) cell growth control in yeast and mammalian cells with autophagy gene deletions. Together, we anticipate that these studies will establish that beclin 1 functions as a tumor suppressor through its Class III-PI3K-dependent autophagy function, and that autophagy represents a fundamental mechanism involved in tumor suppression and negative growth control.

描述（由申请人提供）：自噬的溶酶体途径是降解长期寿命的细胞蛋白和细胞质细胞器的主要调节分解代谢机制。然而，直到最近，对自噬的分子机制或生物学功能知之甚少。在该奖项的资助下，我们的实验室确定了第一个哺乳动物自噬基因Beclin 1（附录项目＃1），并证明它是一个充满单倍抑制的肿瘤抑制基因（附录项目＃2）。我们还发现，Beclin 1和其他自噬基因对于秀丽隐杆线虫中的发育停滞表型至关重要，而秀丽隐杆线虫则受到哺乳动物致癌I类PI3激酶途径的线虫对等效的负面调节（附录项目＃3）。这些发现共同提出了自噬作为肿瘤抑制机制的强烈可能性。在此续订应用程序中，我们将在以下两个具体目标中评估这一概念。在第一个目的中，我们将测试以下假设：Beclin 1通过涉及诱导自噬的III类PI3激酶依赖机制发挥肿瘤抑制活性。为了实现这一目标，我们将使用先前建立的人MCF7乳腺癌细胞模型来确定Beclin 1结合伴侣III PI3-K/HVPS34（涉及自噬囊泡成核）以及自噬基因是否涉及自噬的自噬（自噬效应）是否涉及自动化效应和完成的作用。 Beclin 1-VPS34在体内肿瘤发生中结合的重要性，通过研究一只包含beclin 1的VPS34结合有缺陷的突变体中的敲入小鼠中的肿瘤发生。在第二个目的中，我们将测试Beclin 1的自噬基因的假说，即Beclin 1的自噬基因是负面生长控制和抑制肿瘤的抑制基因所必需的。为了实现这一目标，我们将研究（1）杂合缺失ATG3，ATG5和ATG7的小鼠的自发性肿瘤发生；（2）具有自噬基因，ATG3，ATG5和Beclin 1的纯合缺失的胚胎干（ES）细胞的体内肿瘤性，以及（3）具有自噬基因缺失的酵母和哺乳动物细胞中细胞生长控制。我们共同预测，这些研究将确定Beclin 1通过其III-PI3K依赖性自噬功能起作用，作为肿瘤抑制剂，并且自噬代表了参与肿瘤抑制和阴性生长控制的基本机制。