Candidate Gene Analysis for Macronutrient Selection QTL

描述（由申请人提供）：本申请是我们先前授权的直接延续，该授权针对控制C67 BL/6 J和CAST/Ei小鼠近交系中大量营养素选择的QTL的遗传作图，这两种小鼠近交系在大量营养素饮食的自选摄入方面明显不同。全基因组扫描显示，在6、8、17、18号染色体和X染色体上，代表自我选择的脂肪和碳水化合物摄入量的性状存在显着连锁。当体重作为协变量时，染色体17和18上的位点也与总能量摄入有关。这是第一个在小鼠中定位的食物偏好或总能量摄入的QTL。目前建议的总体目标是确定潜在的候选基因Mnif 1，位于染色体8上的膳食脂肪摄入量的数量性状位点（QTL），和染色体17上的碳水化合物和千卡摄入量的Mnic 1。在目标1中，我们将通过开发B6.CAST同源和亚同源系，分离并缩小Mnif 1和Mnic 1间隔至适合位置候选方法的大小。在目标2中，基因表达微阵列将用于鉴定候选基因。微阵列的探针将由来自受体菌株（B6）和我们实验室开发的间隔特异性亚同源菌株（B6.CAST）的组织的cDNA组成，mRNA将从在食物摄取表型调节中重要的组织（即下丘脑、孤束核/最后区、肝、胃、小肠、胰腺、脂肪细胞、肌肉）中收获。首先，将使用小鼠寡核苷酸阵列（来自UniGene的16，463个基因）进行全基因组基因表达筛选，该阵列现已准备就绪并可从PBRC Genomics Core获得。因此，我们将描述发生在QTL内的基因效应的上游或下游的转录差异。这些数据将提供一个切入点，用于模拟这些喂养行为的控制过程，并确定最有前途的组织与QTL特异性阵列的轮廓。接下来，定制阵列将被设计用于对存在于亚同源间隔中的所有小鼠和人类基因的转录活性进行全面分析，所述亚同源间隔用于控制Chr 8（Mnif 1）上的脂肪摄入和Chr 17（Mnic 1）上的碳水化合物摄入的QTL。饲养性状的生物学知识和QTL内外差异表达基因的分析将帮助我们将候选基因的数量减少到非常少，并选择那些值得进一步研究的功能作用，在确定表型。候选基因的鉴定将增强我们对食物摄入调节的理解。在小鼠中发现调节大量营养素摄入的基因将有助于我们了解影响人类食物偏好的遗传因素与环境因素的作用。这将导致对肥胖和糖尿病的有价值的见解，以及修改宏量营养素选择的新方法，这些方法可能有助于控制体重增加或促进体重减轻。

DESCRIPTION (provided by applicant): This application is a direct continuation of our previous grant directed at the genetic mapping of QTL controlling macronutrient selection in the C67BL/6J and CAST/Ei mouse inbred strains which differ markedly in their self-selected intake of macronutrient diets. A genome-wide scan revealed significant linkage for traits representing self-selected fat and carbohydrate intake on chromosome 6, 8, 17, 18 and X. Loci on chromosome 17 and 18 were linked also to total energy intake when body weight was used as a covariate. These are the first QTL for food preference or total energy intake that have been mapped in the mouse. The overall goal of the current proposal is to identify candidate genes underlying Mnif1, a quantitative trait locus (QTL) for dietary fat intake located on chromosome 8, and Mnic1 on chromosome 17 for carbohydrate and kilocalorie intake. In Aim 1, we will isolate and narrow the Mnif1 and Mnic1 intervals to a size suitable for the positional candidate approach, by developing B6.CAST congenic and subcongenic lines. In Aim 2, gene expression microarrays will be used to identify candidate genes. The probes for the microarrays will consist of cDNA from tissues of the recipient strain (B6) and the interval-specific subcongenic strains (B6.CAST) developed in our laboratory, mRNA will be harvested from tissues important in the regulation of food intake phenotypes, i.e, hypothalamus, solitary tract nucleus/area postrema, liver, stomach, small intestine, pancreas, adipocyte, muscle. First, a genome-wide gene expression screen will be performed using a mouse oligonucleotide array (16,463 genes from UniGene) now ready and available from the PBRC Genomics Core. Thus we will characterize transcriptional differences that occur upstream or downstream from effects of genes within the QTL. These data will provide an entry point for modelling the process by which these feeding behaviors are controlled, and for identifying the most promising tissues to profile with the QTL-specific arrays. Next, custom arrays will be designed for the purpose of performing comprehensive analyses of the transcriptional activity of all mouse and human genes present in the subcongenic intervals for the QTL controlling fat intake on Chr 8 (Mnif1) and carbohydrate intake on Chr 17 (Mnic1). Knowledge of the biology of the feeding traits and analysis of differentially expressed genes within and outside of the QTL will help us reduce the number of candidates to a very few and select those that deserve further investigation for a functional role in determining the phenotype. Candidate gene identification will enhance our understanding of the regulation of food intake. Finding genes regulating macronutrient intake in mice will help us to understand the contribution of genetic versus environmental factors affecting food preferences in humans. This should lead to valuable insights into obesity and diabetes, and new approaches for modifying macronutrient selection that could be useful in controlling weight gain or promoting weight loss.