Investigating the sex-specific roles of PKLR and CHCHD6 in insulin resistance and NAFLD

项目摘要/摘要 非酒精性脂肪性肝病(NAFLD)越来越被认为是酒精性脂肪性肝病的肝脏表现 可导致心血管疾病的代谢综合征。目前，人们对基因的了解很少 以及NAFLD背后的分子因素。此外，目前还没有已知的治疗策略可用于预防或 治疗这种情况。为了解决这个问题，我们应用了一种综合的多组学方法，使用了广泛的 表型小鼠队列，杂交小鼠多样性小组(HMDP)，并确定了许多已知和几个新的 调控NAFLD的候选基因。我们进一步验证了两个新的候选者，丙酮酸激酶，肝脏亚型 (PKLR)和包含6的螺旋结构域(CHCHD6)。当这两个基因 在脂肪变性的雄性小鼠模型中，NAFLD和胰岛素敏感性都得到了改善，通过 肝内甘油三酯蓄积水平，葡萄糖和胰岛素耐量试验，以及空腹血糖和 胰岛素水平。后续的生物能量学分析显示，这两个基因调控着线粒体的代谢。 出现脂肪变性和胰岛素抵抗。当我们进一步扩大到包括女性HMDP菌株时， 我们发现这两个基因没有任何作用。具体地说，相关分析显示PKLR和CHCHD6 不要在雌性品系中分别出现NAFLD和胰岛素抵抗。为了加深对……的理解 这两个基因在调节线粒体代谢中介导的基因与性别的相互作用 在NAFLD和胰岛素抵抗的发展过程中，我提出了三个相互关联的目标来研究1)机制 潜在的PKLR和CHCHD6介导的脂代谢，2)肝脏特异性的代谢后果 雌性小鼠脂肪变性模型中的过度表达或基因敲除，3)基因网络和细胞异质性 在晚期NASH和纤维化的小鼠模型中，肝脏特异性过度表达或敲除改变。 此外，我还制定了详细的五年职业发展计划，其中包括两年 指导阶段(K99阶段)，然后是为期三年的独立阶段(R00阶段)，用于过渡到 独立的学术生涯。我已经包括了一个非常有才华和杰出的顾问小组，包括 Aldons J.Lusis、Orian Shirihai、Neil Kaplowitz、Andrea Hevener、Peter Tontonoz和Steven Bensinger博士 帮助我过渡到独立。我在研究宿主遗传学和性行为方面有丰富的经验 使用系统遗传学分析，差异决定了复杂疾病特征的结果。我现在计划 扩大我在肝脏和线粒体生理学、脂代谢、代谢表型和单细胞方面的培训 测序。总体而言，我的目标是提高我的培训，成为一名独立的调查员，并执行 研究连接系统遗传学、线粒体和肝脏新陈代谢。

PROJECT SUMMARY/ABSTRACT Non-alcoholic fatty liver disease (NAFLD) is increasingly being recognized as the hepatic manifestation of metabolic syndrome that can lead to cardiovascular diseases. Currently, very little is known about the genetic and molecular factors underlying NAFLD. Besides, no known therapeutic strategies are available to prevent or treat the condition. To address this problem, we applied an integrative multi-omics approach using an extensively phenotyped mouse cohort, Hybrid Mouse Diversity Panel (HMDP) and identified many known and several novel candidate genes regulating NAFLD. We further validated two novel candidates, pyruvate kinase, liver isoform (PKLR) and coiled-coil-helix-coiled-coil-helix domain containing 6 (CHCHD6). When these two genes were knockdown in male mouse models of steatosis, both NAFLD and insulin sensitivity improved as measured by intrahepatic triglyceride accumulation levels, glucose- and insulin- tolerance tests, and fasting glucose and insulin levels. Follow-up bioenergetic analyses revealed these two genes regulated mitochondrial metabolism in developing steatosis and insulin resistance. When we further expanded this to include the female HMDP strains, we found that these two genes have no effect. Specifically, correlation analyses revealed PKLR and CHCHD6 do not develop NAFLD and insulin resistance in female strains, respectively. To further our understanding on the gene-by-sex interactions mediated by these two genes in regulating mitochondrial metabolism and in developing NAFLD and insulin resistance, I have proposed three interrelated aims to examine 1) mechanisms underlying PKLR- and CHCHD6- mediated lipid metabolism, 2) metabolic consequences of liver-specific overexpression or knockdown in female mouse models of steatosis, 3) gene networks and cellular heterogeneity altered by liver-specific overexpression or knockdown in mouse models of advanced NASH and fibrosis. Furthermore, I have developed a detailed five-year career development plan that includes two-year mentored phase (K99 phase) followed by three-year independent phase (R00 phase) for my transition into an independent academic career. I have included a highly talented and distinguished panel of advisors including Drs. Aldons J. Lusis, Orian Shirihai, Neil Kaplowitz, Andrea Hevener, Peter Tontonoz, and Steven Bensinger, to assist in my transition to independence. I have significant experience in studying how host genetics and sex differences dictate the outcomes of complex disease traits using systems genetics analyses. I now plan to expand my training in liver and mitochondrial physiology, lipid metabolism, metabolic phenotyping and single cell sequencing. Overall, the goal is to advance my training to become an independent investigator and perform research bridging systems genetics, mitochondria and liver metabolism.