Adaptation to long chain fatty acid oxidation deficiency

描述（由申请人提供）：超长链acyI-CoA脱氢酶（VLCAD）催化哺乳动物线粒体脂肪酸氧化（FAO）螺旋的第一步，在心脏和其他高氧化组织中产生能量。人类隐性VLCAD缺陷产生早期致命性心肌病和心律失常、婴儿低酮性低血糖和肝功能衰竭、婴儿猝死（SIDS）和后来的发作性骨骼肌病的表型。我们的第一个假设是，VLCAD缺乏具有基因型-表型相关性，我们将通过分子遗传学研究（Aim 1），与临床表型相关，并通过研究一些VLCAD突变的病理遗传学，特别是通过新生儿筛查发现的患者中存在的V243A。我们的第二个假设是，VLCAD对心脏功能至关重要，不仅因为它的酶促作用，还因为VLCAD是多种基因转录调控所必需的。这将通过表征我们分离的20 bp长的VLCAD增强子（Aim 2）和表征VLCAD缺陷小鼠及其杂合子幼崽来研究，这些幼崽表现出猝死、迟发性肥胖、肿瘤发展和诱发性心律失常等表型，并因禁食和暴露于寒冷而死亡。该表征包括使用转基因小鼠和细胞转染来检测VLCAD转录调节（目标2）和遥测、侵入性电生理学、微阵列分析、运动测试和VLCAD -/-和+/-动物的饮食操纵（目标3）。我们还将创建组织特异性（心脏、棕色脂肪、肝脏、肌肉）基因敲除（目标4），以确定这些器官对表型的相对贡献。我们知道VLCAD -/-动物在一天内表现出明显的组织基因表达改变，这是围产期的“适应性反应”。我们假设这种挽救生命的适应性反应在基因表达上产生了差异，这些差异是猝死、心律失常、肥胖和肿瘤发生等后期病理的基础。因此，这是一个模型，说明一个基因的多态性或突变如何改变生命后期其他基因的表达，从而导致病理。在这种遗传反应中，一个高度诱导的基因是一种新的蛋白质，称为心脏内皮素相关物质（HERS），基于同源性和表达数据，我们假设其在心脏发育和钙代谢中起作用。我们将在最后一个特定目标中通过产生HERS基因敲除来研究其作用。

DESCRIPTION (provided by applicant): Very-long-chain acyI-CoA dehydrogenase (VLCAD) catalyzes the first step of the mammalian mitochondrial fatty acid oxidation (FAO) spiral that generates energy in the heart and other highly oxidative tissues. Human recessive VLCAD deficiency produces phenotypes of early fatal cardiomyopathy and arrhythmias, infantile hypoketotic hypoglycemia and liver failure, sudden death in infancy (SIDS), and later episodic skeletal myopathy. Our first hypothesis is that VLCAD deficiency has a genotype-phenotype correlation that we will investigate (Aim 1) through molecular genetics, correlating with clinical phenotypes, and by studying the pathogenetics of some VLCAD mutations, particularly V243A that exists in patients discovered by newborn screening. Our second hypothesis is that VLCAD is essential for cardiac function both because of its enzymatic role, but also because VLCAD is required for transcriptional regulation of multiple genes. This will be investigated by characterizing the 20 bp long VLCAD enhancer we have isolated (Aim 2) and by characterizing VLCAD-deficient mice and their heterozygote littermates that exhibit the phenotypes of sudden death, late-onset obesity, development of tumors, and inducible arrhythmias and that die in response to fasting and exposure to cold. This characterization includes use of transgenic mice and cell transfections to examine VLCAD transcriptional regulation (Aim 2) and telemetry, invasive electrophysiology, microarray analysis, exercise testing, and dietary manipulation of VLCAD -/- and +/- animals (Aim 3). We will also create tissue-specific (heart, brown fat, liver, muscle) knockouts (Aim 4) to ascertain the relative contributions of these organs to phenotypes. We know that VLCAD -/- animals exhibit marked alterations in tissue gene expression by one day, a perinatal "adaptive response". We hypothesize that this live-saving adaptive response creates differences in gene expression that are the substrate for the later pathologies of sudden death, arrhythmias, obesity, and tumorigenesis. This is, thus, a model of how polymorphisms or mutations in one gene modify expression of other later in life, leading to pathology. One highly induced gene in this genetic response is a novel protein, designated heart endothelin-related substance (HERS) that, based upon homology and expression data, we postulate functions in cardiac development and calcium metabolism. We will investigate its role by generating a HERS knockout in the last specific aim.