Regulation of energy homeostasis and adiposity by a Golgi-associated PARP enzyme

描述(由申请人提供)： 项目摘要许多退伍军人患有肥胖症，这是一种由于能量摄入超过能量消耗而导致的疾病。为了解决这一健康问题，重要的是要确定新的能量平衡调节器，这些调节器可以接受药物干预。我们的长期目标是阐明Tankyrase(TNKS)是如何调节能量平衡的，TnKS是一种高尔基体相关酶，通过多聚ADP核糖化来修饰底物蛋白。这一应用的目的是研究tankyrase在调节能量消耗中的作用，重点是燃料底物在器官之间的分配以及脂肪细胞脂联素的分泌。为此，我们在小鼠中使用了基因捕获技术来建立Tankyrase缺陷(TNKS-/-)模型。我们发现，与野生型对照相比，TNKS-/-小鼠增加了能量消耗，减少了肥胖，并增加了食物摄入量。有趣的是，他们的血浆脂联素水平，特别是具有生物活性的高分子形式，正在强劲地增加。与已知的脂联素作用一致，TNKS-/-小鼠的肌肉显示出介导甘油三酯和脂肪酸分解代谢的基因上调。另一方面，这些小鼠的肝脏显示出促进甘油三酯释放的基因上调。基于这些和其他观察，我们假设，TNKS-/-小鼠能量消耗的增加导致肝脏甘油三酯分泌增加，甘油三酯被输送到肌肉进行分解代谢，以响应高脂联素血症。在这个工作模型中，从肝脏到肌肉的脂质流动是以牺牲脂肪组织中的储存为代价的，导致TNKS-/-小鼠的瘦表型，尽管食物摄入量有了代偿性增加。为了验证我们的工作假设，目标1将确定TNKS缺乏在多大程度上增加了体内循环甘油三酯的产生和清除，以及肌肉在体外水解甘油三酯和氧化脂肪酸的活性。目的2将评估这样的预测，即TNKS-/-小鼠由于能量消耗增加，在挑战高脂肪饮食时可免受肥胖和脂肪毒性的影响。目的3研究TnKS在脂联素分泌和能量平衡中的脂肪特异性作用。初步研究表明，TNKS-/-小鼠的高脂联素血症是一种转录后效应，可以通过敲除3T3-L1脂肪细胞中的TNKS来重现。我们将对3T3-L1脂肪细胞应用[35S]脉冲追逐分析，以测试TNKS基因敲除在分泌途径中稳定脂联素的预测。我们还将使用条件基因敲除方法来确定脂肪特异性消融TNKS在多大程度上重现了TNKS-/-小鼠的高脂联素血症和其他代谢表现。这项拟议的研究有望阐明TNKS在脂联素分泌和能量平衡中的作用，以及燃料底物在分解代谢和储存之间的分配。这一应用的结果可能为进一步的研究提供基础，以验证TNKS作为抗肥胖治疗的药理靶点。 公共卫生相关性： 项目简介超过能量消耗的饮食卡路里摄入量作为脂肪储存在脂肪组织中。过量摄入卡路里的长期后果是肥胖，这是2型糖尿病、心脏病发作和其他严重并发症的主要风险因素。根据最近的一项调查，在退伍军人管理局医疗机构服务的人群中，肥胖影响多达37%的女性和33%的男性。因此，改善肥胖对许多退伍军人的健康非常重要。使用小鼠模型来探索肥胖的调节，我们观察到当一种名为tankyrase的细胞蛋白失活时，出现了一种瘦肉表型。我们的数据表明，这些缺乏tankyrase的小鼠因为能量消耗增加而变瘦。我们建议研究缺乏tankyrase功能导致能量消耗增加的分子机制。我们的长期目标是建立tankyrase作为治疗肥胖症的药理靶点。

DESCRIPTION (provided by applicant): Project Summary Many veterans suffer from obesity, an illness that results from a cumulative excess of energy intake over energy expenditure. To combat this health issue, it is important to identify novel regulators of energy homeostasis that are amenable to pharmacological intervention. Our long-term goal is to elucidate how energy homeostasis is regulated by tankyrase (TNKS), a Golgi-associated enzyme that modifies substrate proteins through poly-ADP-ribosylation. The aim of this application is to investigate the role of tankyrase in regulating energy expenditure with an emphasis on fuel substrate partitioning between organs as well as adiponectin secretion from adipocytes. To this end, we have used the gene- trapping technique in mice to create a tankyrase deficient (TNKS -/-) model. We found that TNKS -/- mice have increased energy expenditure, reduced adiposity, and greater food intake than wild-type controls. Intriguingly, their plasma levels of adiponectin, particularly the bioactive high molecular-weight form, are robustly increased. Consistent with the known effect of adiponectin, muscle in TNKS -/- mice shows an upregulation of genes that mediate the catabolism of triglycerides and fatty acids. On the other hand, liver in these mice shows an upregulation of genes that promote triglyceride release. Based on these and other observations, we hypothesize that the heightened energy expenditure in TNKS -/- mice entails increased hepatic secretion of triglycerides, which are channeled to the muscle for catabolism in response to hyperadiponectinemia. In this working model, the lipid flux from liver to muscle is at the expense of storage in adipose tissue, resulting in the lean phenotype of TNKS -/- mice despite a compensatory increase in food intake. To test our working hypothesis, Aim 1 will determine the extent to which TNKS deficiency increases the production and clearance of circulating triglycerides in vivo as well as the activity of muscle to hydrolyze triglycerides and oxidize fatty acids ex vivo. Aim 2 will evaluate the prediction that TNKS -/- mice, owing to increased energy expenditure, are protected from obesity and lipotoxicity when challenged with a high fat diet. Aim 3 will investigate the adipose-specific roles of TNKS in adiponectin secretion and energy homeostasis. Preliminary studies have shown that the hyperadiponectinemia of TNKS -/- mice is a post-transcriptional effect and can be recapitulated by knocking down TNKS in 3T3-L1 adipocytes. We will apply [35S] pulse-chase analysis to 3T3-L1 adipocytes to test the prediction that TNKS knockdown stabilizes adiponectin in the secretory pathway. We will also use the conditional knockout approach to determine the extent to which adipose-specific ablation of TNKS recapitulates the hyperadiponectinemia and other metabolic manifestations of TNKS -/- mice. The proposed studies are expected to shed light on the roles of TNKS in adiponectin secretion and energy homeostasis as well as the partitioning of fuel substrates between catabolism and storage. Outcomes from this application may provide the foundation for additional studies that validate TNKS as a pharmacological target for anti-obesity therapy. PUBLIC HEALTH RELEVANCE: Project Narrative Dietary calorie intake that exceeds energy expenditure is stored in the adipose tissue as fat. A long-term consequence of excessive caloric intake is obesity, which is a major risk factor for type 2 diabetes, heart attack and other serious complications. According to a recent survey, obesity affects as many as 37% of women and 33% of men among the population served by VA medical facilities. Therefore, amelioration of obesity is important to the health of many veterans. Using a mouse model to explore the regulation of adiposity, we have observed a lean phenotype when a cellular protein called tankyrase is inactivated. Our data indicate that these tankyrase-deficient mice are lean because of heightened energy expenditure. We propose to investigate the molecular mechanisms whereby lack of tankyrase function leads to heightened energy expenditure. Our long-term goal is to establish tankyrase as a pharmacological target in the treatment of obesity.