Elucidating a mechanism of mTORC1 activation independent of amino acids signaling

阐明独立于氨基酸信号传导的 mTORC1 激活机制

• 批准号: 8443550
• 负责人: David M. Sabatini
• 金额: $ 29.25万
• 依托单位: WHITEHEAD INSTITUTE FOR BIOMEDICAL RES
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-09-30 至 2014-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8443550
• 关键词: Address; Adenoviruses; Affect; Aging; Albumins; Alleles; Amino Acids; Amino Acids Activation; Autophagocytosis; Biogenesis; Breeding; Calnexin; Candidate Disease Gene; Categories; Cell Line; Cells; Complementary DNA; Diabetes Mellitus; Disease; Dominant-Negative Mutation; Down-Regulation; Early Endosome; Excision; Fasting; Gene Expression Profile; Genes; Genetically Engineered Mouse; Golgi Apparatus; Growth; Growth Factor; Guanosine Triphosphate Phosphohydrolases; Hepatocyte; Immunofluorescence Immunologic; Immunoprecipitation; Infection; Institutes; Libraries; Liver; Lysosomes; Malignant Neoplasms; Mass Spectrum Analysis; Mediating; Membrane; Membrane Protein Traffic; Messenger RNA; Molecular; Mus; Null Lymphocytes; Nutrient; Organism; PTEN gene; Pathway interactions; Phenotype; Phosphoric Monoester Hydrolases; Phosphorylation; Phosphotransferases; Physiology; Protein Family; Protein Kinase; Protein Phosphatase 2A Regulatory Subunit PR53; Protein phosphatase; Proteins; Proteomics; RNA Interference; Raptors; Regulation; Resistance; Series; Signal Pathway; Signal Transduction; Sirolimus; Staining method; Stains; Structure; Subfamily lentivirinae; Surface; System; TSC1/2 gene; Technology; Transgenic Mice; Tubulin; Virus; Withdrawal; Work; age related; base; candidate validation; cell growth; cost; deprivation; desensitization; embryo cell; falls; feeding; follow-up; genome-wide; human disease; immortalized cell; improved; in vivo; inhibitor/antagonist; knock-down; liver infection; mouse model; novel; novel therapeutic intervention; novel therapeutics; overexpression; research study; small hairpin RNA; stable cell line

DESCRIPTION (provided by applicant): The mechanisms underlying the pro-aging consequences of mTOR activity are obscure. Deregulated mTOR activity leads to accelerated aging and upregulated mTORC1 activity occurs in aging organisms. We recently described the Rag family of proteins as key for mTORC1 activation by amino acids, through a mechanism that requires mTORC1 shuttling to the lysosomal surface. We have generated a series of immortalized cell lines derived from genetically-engineered mice that lack the RagA gene. Although mTORC1 activity is barely detectable in RagA-deficient embryos, cell lines derived from them have reactivated mTORC1 signaling, which is now insensitive to amino acids withdrawal, but sensitive to growth factors withdrawal. Intriguingly, in these lines mTORC1, although active, does not localize to lysosomes, indicating that an additional unknown mechanism of mTORC1 activation is at work. Based on a combination of transcriptional profiling and proteomic-based approaches, we plan to find the genes responsible for the lysosomal-independent activation of mTORC1 and to understand how this occurs. We will then take advantage of RagA-null livers to study in vivo the consequences of altering the expression of the candidates found. Further elucidation of the molecular mechanism leading to mTORC1 desensitization to amino acid withdrawal, together with the identification of druggable targets within this pathway, may pave the way for novel therapeutic approaches to aging and age-related diseases. PUBLIC HEALTH RELEVANCE: Deregulated cell growth signals arising from the mechanistic target of rapamycin (mTOR) protein kinase occur in diabetes, cancer and aging. Thus, understanding mTOR signaling pathway will enable the pursuit of improved therapies against these diseases. Taking advantage of novel mouse models of deregulated mTOR activity we have generated cell lines with a unique and intriguing regulation of mTOR activation, resistant to nutrient deprivation, which normally inactivates this signaling pathway. The identification of the responsible genes and the molecular mechanisms governing this particular signaling alteration that we plan to investigate in the present proposal will help understand deregulated cell growth states. The series of novel mouse models will also allow us to explore the consequences of this deregulated growth state in mammalian physiology, helping us to develop novel therapeutic avenues to manipulate the mTORC1 pathway in human disease.

描述（由申请人提供）：mTOR 活性促衰老后果的机制尚不清楚。 mTOR 活性失调会导致衰老加速，而衰老生物体中 mTORC1 活性上调。我们最近将 Rag 蛋白家族描述为氨基酸激活 mTORC1 的关键，其机制需要 mTORC1 穿梭到溶酶体表面。我们已经产生了一系列源自缺乏 RagA 基因的基因工程小鼠的永生化细胞系。尽管在 RagA 缺陷胚胎中几乎检测不到 mTORC1 活性，但源自它们的细胞系已重新激活 mTORC1 信号传导，该信号现在对氨基酸撤退不敏感，但对生长因子撤退敏感。有趣的是，在这些细胞系中，mTORC1 虽然有活性，但并不定位于溶酶体，这表明 mTORC1 激活的另一个未知机制正在发挥作用。基于转录分析和基于蛋白质组学的方法的结合，我们计划找到负责 mTORC1 溶酶体独立激活的基因，并了解这是如何发生的。然后，我们将利用 RagA 无效的肝脏来研究体内改变所发现的候选者表达的后果。进一步阐明导致 mTORC1 对氨基酸戒断脱敏的分子机制，以及该途径中可药物靶点的鉴定，可能为衰老和年龄相关疾病的新治疗方法铺平道路。 公共健康相关性：雷帕霉素 (mTOR) 蛋白激酶机制靶点产生的细胞生长信号失调发生在糖尿病、癌症和衰老过程中。因此，了解 mTOR 信号通路将有助于寻求针对这些疾病的改进疗法。利用 mTOR 活性失调的新型小鼠模型，我们生成了具有独特且有趣的 mTOR 激活调节的细胞系，能够抵抗营养剥夺（营养剥夺通常会使该信号通路失活）。我们计划在本提案中研究的负责基因和控制这种特定信号改变的分子机制的识别将有助于了解失调的细胞生长状态。这一系列新型小鼠模型还将使我们能够探索这种失调的生长状态对哺乳动物生理学的影响，帮助我们开发新的治疗途径来操纵人类疾病中的 mTORC1 通路。