线粒体相关基因遗传变异与麻风易感性关联分析

Leprosy is a chronic, systemic disease caused by Mycobacterium leprae. It affects human skin and the peripheral nerve system, as well as eyes and other systems, resulting in a variety of disabilities associated with leprosy. Leprosy control in China, particularly Yunnan Province, is still urgent, despite the fact that we had successfully achieved the eliminate goal at country level during the past decades. As an ancient disease, leprosy can be used as a good model to study the innate immune response to pathogen and to uncover the interaction between immune system and nerves system during the onset of leprosy. Genetic factors confer susceptibility to this disease. During the long evolution, more than half of the functional genes in the genome of M. leprae are inactivated or have became pseudogenes, and this bacterium has developed a dependence on host energy production and nutritional products. As a result, the parasitic life and adaptation might have shaped host genetic susceptibility to leprosy. In this study, we analyze genetic polymorphisms of 9 genes (LRRK2、PINK1、PARL、POLG、TFAM、PEO1、MFN2、MAVS and MITA) that are actively involved in mitochondrial function. We choose a sufficient number of tag SNPs to cover each gene and use a high throughput genotyping technology to genotype each SNP. Associations of alleles, genotypes, and haplotypes of all genes with leprosy and its subtypes are evaluated. We further validate the functional SNPs at the cellular and molecular levels. This study will provide essential information for us to understand the role of mitochondrial genes confering genetic susceptibility to leprosy. Meanwhile, our results will have implications for us to study the immuno-neurological interaction in other infectious disease and neurodegenerative diseases.

麻风是由麻风杆菌引起的一种慢性传染病，对病人的皮肤、外周神经伤害极大，严重可致残和致盲。麻风防治工作在我国（尤其是云南）仍相当紧迫。麻风易感性和临床表型受到宿主遗传因素影响，但其中起关键作用的基因及相应机理尚未完全清楚。在进化过程中麻风杆菌大量基因功能丢失，对宿主的能量代谢系统产生严重依赖，宿主能量代谢方面的差异可能影响其易感。本项目将集中对9个线粒体相关基因（LRRK2、PINK1、PARL、POLG、TFAM、PEO1、MFN2、MAVS和MITA）遗传变异进行高通量基因分型，分析基因型、单倍型与临床表型间的关联性，以期确定易感遗传变异，并从分子和细胞水平上对易感基因开展功能验证实验，探讨其调控机制及在麻风杆菌感染过程中可能发挥的作用。结果有望阐明线粒体相关基因遗传变异与麻风遗传易感性之间的关系，提供该病防治的新思路，同时为其它细菌感染性疾病和神经退行性疾病的研究提供参考和启示。

本项目基于麻风临床特征及其病原麻风分枝杆菌对于宿主能量代谢系统极度依赖的特性，围绕“线粒体相关基因遗传变异与麻风易感”这一主题开展麻风的遗传分析。通过对云南玉溪和文山地区麻风病人开展系统的遗传分析，取得以下发现：（1）线粒体外膜蛋白LRRK2上错义突变p.M2397T影响麻风的易感性，功能实验揭示p.M2397T对NFAT的激活效应明显，且能抑制由神经毒素MPTP诱导的自噬；（2）线粒体内膜蛋白OPA1、PARL和PINK1基因变异与麻风，尤其是多菌型麻风显著相关。蛋白互作网络分析显示，OPA1、PARL和PINK1直接或者间接与报道的麻风易感基因、多个神经和免疫系统基因相互作用；（3）线粒体复制相关蛋白TFAM、PEO1和POLG与麻风的关联研究发现，TFAM和POLG的基因变异能够影响麻风的遗传易感，相关结果正整理成稿中；（4）线粒体先天免疫蛋白MAVS、MITA和MFN2的遗传变异与麻风易感性没有相关性，提示线粒体先天免疫相关基因是否参与麻风易感需要更多的研究来确证；（5）我们富集了719个线粒体相关基因，设计和优化了基于二代测序技术的目标基因靶向捕获技术，并在文山样本中开展了深度测序，目前数据正在分析中，有望成功描绘影响麻风易感的线粒体基因致病突变谱；（6）补体基因FCN2、MBL2和CFH的遗传变异影响麻风易感，且FCN2和MBL2低转录活性的启动子单倍型是少菌型麻风的风险因素；（7）对目前所有已报道的麻风相关基因进行了一个系统的整合分析，获得123个与麻风相关且同时在患者皮损组织中表达有改变的基因，这些基因显著地富集在自身免疫疾病相关通路，提示麻风可能是一种自身免疫病；（8）成功地验证到前人全基因组关联分析报道的rs3762318位点与麻风相关，进一步eQTL、组织表达谱和蛋白互作网络等分析发现，该位点可能影响SLC35D1基因在麻风皮损组织中的表达，提示SLC35D1是麻风潜在易感基因。以上研究结果大大地丰富了我们对于麻风遗传易感的认识，显示线粒体相关基因遗传变异与麻风遗传易感性存在较大关联，支持我们基于麻风分枝杆菌由于基因组丢失，导致对宿主能量代谢系统依赖而提出的宿主线粒体基因变异影响麻风遗传易感性的假说，同时为其它细菌感染性疾病和神经退行性疾病的研究提供了参考和启示。项目执行期间共发表SCI 论文8篇。培养毕业硕士研究生3名，博士研究生2名。