MUTYH功能缺陷介导线粒体非折叠蛋白应答与衰老及相关疾病表型发展的分子基础研究

With social development, aging and age-related diseases have become the biggest medical problem in threatening health and life of human beings. It is very important for us to investigate the complicated biological process of aging and clarify the pathogenesis of aging-related disease. Recent studies suggested that mitochondrial stress induced with oxidative damage, mitochondrial unfolded protein response (UPRmt) and the progressive development of mitochondrial function impairment is the basic pathophysiological process of aging and age-related diseases, but its molecular mechanism has not been fully elucidated still. The MUTYH is involved in the base excision repair system, which is one of the critical genes for repairing oxidized DNA bases and maintaining the fidelity of DNA replication. We had previously described a novel variant of AluYb8 insertion in the MUTYH genes, which would induced a selective inhibition expression of MUTYH 1 protein that localized at mitochondria, in the carriers with the homozygous mutation. It caused mtDNA content decreased and mitochondrial function weakened, and resulted in mitochondrial stress. Interestingly, the mtDNA content was increased in the patients of age-related diseases with the MUTYH homozygous mutation, but the expression of their mtDNA genes decreased. It indicated an mitochondrial unfolded protein response disorder. A similar result was also found in the pulmonary fibrosis model of mice with the MUTYH knockout. We considered that the characteristic change of mitochondrial DNA content mediated by MUTYH dysfunction could be taken as an important biomarker for aging and age-related disease. Based on the mentioned above, we propose the project for study on the MUTYH dysfunction-mediated mitochondrial unfolded protein response and the molecular mechanism of aging and age-related diseases. We will use the MUTYH knockout mice to established the aging animal model, and used the cultured primary embryonic mouse cells to set up the senescence cell model. We would like to concentrate on the MUTYH dysfunction, mtDNA alteration, UPRmt, and the genetic and epigenetic regulation of their signaling molecule expression, and to address the pathogensis and key moleculars of the age-related diseases. Implementation of this project could be expected to clarify the phenotype drive modulation of aging and age-related disease and the results of our study could be favorable for the translational medicine of aging and age-related diseases.

目前认为，氧化损伤引起线粒体应激，线粒体非折叠蛋白应答（UPRmt）,线粒体功能障碍渐进发展是衰老及相关疾病的基本病理生理过程，但其分子机制远未阐明。我们前期在人类样本和动物模型的研究中均发现，MUTYH基因缺陷可导致机体氧化应激、线粒体功能受损。在个体衰老和相关疾病表型发生过程中，携带者细胞mtDNA含量呈特征性变化，提示存在UPRmt，并伴有线粒体自噬等相关分子表达改变，其表型意义有待深入研究。基于此，我们提出MUTYH功能缺陷介导UPRmt与衰老表型发展的分子基础研究。我们拟采用MUTYH基因敲除小鼠构建衰老动物模型；同时利用同源胚鼠细胞的原代培养，建立不同暴露因素诱导的体外细胞模型。系统探讨衰老及表型发展中UPRmt相关分子功能和表达的改变、遗传与表观遗传调控机制；深入分析mtDNA特征性改变的表型意义、衰老及相关疾病的关键驱动分子。为发展衰老及相关疾病的临床干预技术探寻潜在靶标。

前期研究发现，MUTYH功能缺陷可导致机体氧化应激、细胞线粒体功能受损，且机体衰老及其关联疾病表型发生过程中，MUTYH缺陷细胞mtDNA呈特征性的改变，提示存在线粒体非折叠蛋白应答（UPRmt）机制。为深入认识其表型意义及其分子基础，本项目利用Mutyh缺陷小鼠及细胞模型，并结合人群样本，系统观察了Mutyh缺陷与线粒体功能障碍、组织细胞病理状态的关系，并分析其分子调控及功能机制。突破性进展主要包括：①MUTYH功能缺陷可引起机体细胞UPRmt，但其产生的病理性影响存在明显的组织学差异，依赖于组织细胞的能量代谢状态；②Mutyh缺陷/氧化应激可抑制线粒体外膜融合蛋白（Mfn1/2）和内膜融合蛋白（Opa1）的表达，严重损害线粒体结构，能量代谢中间分子α-酮戊二酸（α-KG）具有重要的调控作用；③Mutyh缺陷明显缓解Bleomycin诱导的小鼠肺纤维化病理表型，其机制可能与Mutyh缺陷关联的细胞基因组DNA单链断裂减少、肺上皮细胞凋亡降低有关；④病例对照分析显示人类MUTYH基因AluYb8插入变异影响携带者衰老相关疾病的发病年龄。本研究结果深化了对MUTYH功能缺陷病理学意义认识，且为以MUTYH为靶点疾病治疗和诊断的转化研究提供了理论依据。