Eukaryotic genes involved in adult lifespan regulation

#The present invention relates to regulation of lifespan in eukaryotes. More particularly, one aspect of the present invention is directed to methods of assaying for genes, gene products, and genes in pathways controlled by such genes and gene products, using RNAi and microarray analysis, that regulate lifespan (e.g., extend or truncate adult lifespan) in eukaryotes such as invertebrates (e.g., C. elegans), plants, and mammals, e.g., humans. For example, one aspect of the present invention is directed to genes, in particular human genes, encoding components of the mitochondrial respiratory chain and genes encoding glycolysis enzymes, which are involved in lifespan regulation, and genes and gene products in pathways controlled by such genes. Other genes and gene products identified as regulating aging and aging pathways include a gene encoding a GTPase; a transcriptional activator; novel genes: llw-1, llw-2, llw-3, and llw-4; genes encoding cytochrome P450 proteins (involved in steroid biosynthesis); a melatonin synthesis gene; genes encoding insulin and insulin-like peptides; genes encoding heat shock factors; genes encoding catalases; stress-response genes; and metabolic genes. The invention further relates to methods for identifying and using agents, including small molecule chemical compositions, antibodies, antisense nucleic acids, and ribozymes, that regulate, e.g., enhance, adult lifespan via modulation of aging associated proteins; as well as to the use of expression profiles, markers, and compositions in diagnosis and therapy related to lifespan extension, life expectancy, and aging. The present invention also relates to gene therapy involving lifespan associated genes.

#Previously, classic genetic screens have been used to identify genes involved in the C. elegans development. In one example, inhibition of mitochondrial respiratory chain genes such as NADH ubiquinone oxidoreductase and ATP synthase in C. elegans larva was found to impair larval development and cause arrest in the third larval stage (see, e.g., Tsang et al., JBC 276:33240-33246 (2001)). In other examples, classical genetic screens have been used to identify C. elegans genes involved in a variety of processes, including dauer formation, and embryonic development. Some of these genes, for example the daf-2 and daf-16 genes, have been implicated in the regulation of lifespan see, e.g., Kenyon et al., Nature 366:461-464 (1993); Morris et al., Nature 382:536-539 (1996); Kimura et al., Science 277:942-946 (1997); Paradis et al., Genes Dev. 12:2488-2498 (1998); Paradis et al., Genes Dev. 13:1438-1452 (1999); Off & Ruvkun, Mol. Cell 2:886-893 (1998); Guarente & Kenyon, Nature 408:255-262 (2000); Ogg et al., Nature 389:994-999 (1997); and Lin et al., Science 278:1319-1322 (1997)).

#Classical genetic screens are frequently time consuming, both in identification of interesting mutants and in cloning a gene associated with a mutation. Classical genetic screens can include labor intensive backcrosses to eliminate mutations unlinked to the phenotype of interest. Classical genetic screens also may require the extra step of cloning the gene of interest, by complementation of the mutation.

#FIG. 1 shows RNAi of respiratory chain and glycolysis genes increasing lifespan.

#FIG. 2 shows daf-2(e1370) worms treated with dsRNAs of respiratory and glycolytic genes.

#FIG. 3 shows antimycin increases lifespan.

#Double-stranded RNA-mediated interference (RNAi) provides a sequence specific mechanism for inhibiting gene expression in C. elegans (see, e.g., Fire et al., Nature 391:806-811 (1998) and WO 99/32619)). This technique is also useful for functional genomics analysis of C. elegans genes (see, e.g., Fraser et al., Nature 408:325-330 (2000); Kamath et al., Genome Biol. 2:RESEARCH0002 (2000)).

#Surprisingly, RNAi can be used to identify genes and gene products involved in lifespan and aging mechanisms in adults, by feeding bacteria expressing a dsRNA library over long-term time periods to C. elegans and measuring, e.g., mean and median lifespan increase or decrease in adults. Such assays can be used, in addition, to determine if two or more genes function together in a similar aging pathway, to determine if gene function is cell autonomous, and to determine if drug compounds known to alter the aging process function in the same or different pathways. Microarray assays can also be used to identify genes and gene products involved in lifespan and aging (see, e.g., Table 5 and Table 6).

#Microarray and RNAi assays are also useful in combination, e.g., by identifying a gene or gene product involved in lifespan using RNAi and then examining its expression pattern in old and young adults using microarray analysis, or by identifying a differentially expressed gene or gene product involved in lifespan using microarray analysis and then examining the effects of gene or gene product inhibition using RNAi. Microarray analysis can also be performed on animals treated with RNAi of genes identified using microarray analysis.