Isoform-specific function of y-AMPK in the heart.

描述（由申请人提供）：amp活化蛋白激酶（AMPK）是一种丝氨酸/苏氨酸激酶，作为细胞内能量传感器和细胞代谢、生长和死亡的关键调节剂，因此是肥胖、糖尿病、癌症和心血管疾病的有希望的药物靶点。AMPK是一种杂三聚物，由催化1-亚基和调节2-亚基和3-亚基组成，每个亚基有多个同工型。先前的研究表明，应激时心脏中AMPK的激活是一种重要的心脏保护机制。然而，AMPK调控32亚基（由prkag2基因编码）的点突变导致人类心肌病，其特征是心肌肥厚、心律失常和糖原储存。这引起了对未来针对心脏AMPK级联的药物开发的关注。因此，了解prkag2心肌病的发病机制是至关重要的。利用在心脏（TG32N488I）中过表达突变体prkag2 （N488I）的转基因小鼠（TG32N488I）真实再现了prkag2心肌病，我们在之前的资助期内证明了该突变在没有能量缺陷的情况下引起AMPK的异常激活，并且可以通过抑制AMPK活性来挽救心肌病表型。我们进一步证明，在没有能量应激的情况下，葡萄糖摄取和脂肪酸氧化的同时增加导致TG32N488I心脏中外源性葡萄糖的代谢重定向优先进入糖原库。为了确定心肌病表型是否完全归因于糖原储存，我们试图通过特异性靶向肌糖原合成酶的活性，在不改变突变基因表达的情况下挽救糖原储存。我们的初步数据显示，该策略成功地使TG32N488I小鼠的心脏糖原含量正常化，但没有使心脏肥厚表型正常化。因此，我们假设32- AMPK突变导致心肌肥厚与糖原储存无关。此外，由于prkag2突变主要引起心脏表型，它提出了32-AMPK在心脏中的独特作用的问题。这导致我们产生32-AMPK缺失的小鼠模型，以确定3-AMPK的同工型特异性功能。利用这些模型，我们提出：1)验证prkag2突变独立于糖原储存导致心脏肥厚的假设，并确定在营养饱和条件下32-AMPK异常激活刺激心脏肥厚的机制；2)明确32-AMPK信号级联在心脏发育和疾病过程中的组成和功能。

DESCRIPTION (provided by applicant): AMP-activated protein kinase (AMPK) is serine/threonine kinase that serves as an intracellular energy sensor and a key regulator of cell metabolism, growth and death, hence a promising drug target for obesity, diabetes, cancer and cardiovascular diseases. AMPK is a heterotrimeric complex composed of catalytic 1-subunit and regulatory 2- and 3-subunits with multiple isoforms for each subunit. Previous studies have shown that activation of AMPK in the heart during stress is an important cardioprotective mechanism. However, point mutations in the regulatory 32-subunit (encoded by prkag2 gene) of AMPK cause human cardiomyopathy characterized by cardiac hypertrophy, arrhythmias and glycogen storage. This raises concerns of future drug development targeting AMPK cascade in the heart. It is thus critical to understand the disease mechanisms responsible for the prkag2 cardiomyopathy. Using transgenic mice overexpressing mutant prkag2 (N488I) in the heart (TG32N488I) that faithfully recapitulated prkag2 cardiomyopathy, we in the previous funding period demonstrated that the mutation caused aberrant activation of AMPK in the absence of energetic deficit, and the cardiomyopathy phenotype could be rescued by inhibition of AMPK activity. We have further demonstrated that the simultaneous increases in glucose uptake and fatty acid oxidation in the absence of energy stress resulted in metabolic re-routing of exogenous glucose preferentially into glycogen pool in the TG32N488I hearts. To determine whether the cardiomyopathy phenotype is entirely attributable to glycogen storage, we sought to rescue the glycogen storage without changing the mutant gene expression by specifically targeting the activity of muscle glycogen synthase. Our preliminary data showed that this strategy successfully normalized cardiac glycogen content but not the cardiac hypertrophy phenotype in TG32N488I mice. Thus, we hypothesize that 32- AMPK mutation causes cardiac hypertrophy independent of glycogen storage. Moreover, since the prkag2 mutation causes predominantly cardiac phenotype, it raises the question of a unique role of 32-AMPK in the heart. This led us to generate mouse models deficient of 32-AMPK in order to determine the isoform-specific function of 3-AMPK. Using these models we propose 1) to test the hypothesis that the prkag2 mutation causes cardiac hypertrophy independent of glycogen storage and, to determine the mechanisms by which aberrant activation of 32-AMPK under nutrient saturated conditions stimulate cardiac hypertrophy; 2) to define the composition and the function of the 32-AMPK signaling cascade in the heart during cardiac development and diseases.