Comparative Functional Genomics of Longevity Assurance

描述(由申请人提供)：我们建议通过对一组不同预期寿命的重组近交系(RI)线虫和小鼠品系以及长寿突变线虫品系的全球基因表达谱来表征长寿保证基因(LAG)。我们的实验设计--从其可比性中获得力量--旨在识别那些基因(A)其转录水平与RI品系之间以及在突变与野生动物中的长寿命一致相关，以及(B)在多个物种中独立发现这种关联。这种实验方法将需要在生物生物学、遗传学、基因组学和统计学方面的协调努力，而聚集在一起的团队成员反映了这种多样性。我们的具体目标是： 1.对两种12个线虫RI系的寿命进行了定量表征。 将确定现有的6个线虫RI系和6个新分离的线虫RI系的平均寿命和最大寿命；2.检验RI系和线虫突变株的全基因组比较表达分析将识别滞后候选者的假设。将在固定的年龄段和生理年龄对6个RI系和2个线虫突变株的基因表达进行全基因组微阵列分析。我们将使用高灵敏度的Q-PCR分析来验证从微阵列分析中挑选的候选基因的表达水平；3.确定哪些候选基因具有特定的目的2是必要的滞后。利用RNAi技术操纵在线虫中发现的特定目标2中候选基因的子集的表达，将允许为这些基因分配相对权重；4.通过转录图谱识别重组布里吉萨线虫自交系中的滞后候选基因。这一具体目标将(A)使我们能够确定延长预期寿命的“私人”和“公共”机制的特征，以及(B)允许一组独立的数据来支持(或反驳)在具体目标3中使用RNAi产生的那些数据。 5.将分析方法推广到小鼠RI系。在第四年和第五年，我们将应用上面定义的功能基因组方法，以及在前三年改进的统计方法，来分析具有不同寿命的小鼠RI系。

DESCRIPTION (provided by applicant): We propose to characterize longevity assurance genes (LAGs) through global gene expression profiling of a panel of recombinant inbred (RI) nematode and mouse strains of varying life expectancy, as well as long-lived mutant nematode strains. Our experimental design--which derives power from its comparative nature--aims to identify those genes (a) for which transcriptional level is consistently associated with long lifespan across RI lines and in mutant versus wild-type animals, and (b) for which such associations are found independently in multiple species. This experimental approach will require coordinated efforts in organismal biology, genetics, genomics, and statistics, and the team members assembled reflect this diversity. Our specific aims are: 1. To quantitatively characterize the lifespans of twelve nematode RI lines in two species. Mean and maximum lifespans will be confirmed for six existing RI lines of C. elegans and six newly-isolated RI lines of C. briggsae; 2. To test the hypothesis that whole-genome comparative expression analysis of RI lines and mutant strains of C. elegans will identify LAG candidates. Whole-genome microarray analysis of gene expression in six RI lines and two mutant strains of C. elegans will be carried out, at fixed chronological and physiological ages. We will employ highly sensitive Q-PCR analysis to validate the expression levels of candidate genes picked from microarray analysis; 3. To determine which of the candidate genes characterized in Specific Aim 2 are necessary LAGs. The use of RNAi technology to manipulate the expression of a subset of candidate genes discovered in Specific Aim 2 in C. elegans will permit a relative weight to be assigned to these genes; 4. To identify LAG candidates in recombinant inbred lines of C. briggsae via transcriptional profiling. This specific aim will (a) allow us to characterize "private" versus "public" mechanisms for increasing life expectancy, and (b) permit an independent set of data to support (or contradict) those data generated with the use of RNAi in Specific Aim 3. 5. To extend the analytical approach to mouse RI lines. In years four and five we will apply the functional genomic approach defined above, as well as statistical methods refined during the first three years, to the analysis of mouse RI lines with differential lifespans.