Our laboratory has been working on elucidating the mechanisms of cellular senescence in mammals, and understanding its contribution to organismal aging. It is generally accepted that telomere shortening is a crucial contributor of aging in mammalian cells, giving rise to the "telomere hypothesis of aging". This hypothesis is currently being tested in mammals, but recent data suggest telomere structure, and possibly telomere organization, may influence cellular, and possibly organismal, phenotypes. We investigated whether mechanisms of telomere length regulation is conserved in the metazoan nematode Caenorhabditis elegans in connection with aging. We examined seven mutants that have been shown to have an lengthened life span for changes in telomere length. Among the seven mutants examined, the long-lived
clk-2 mutant showed a shortened telomere length compared with wild type N2 worms. Moreover,
clk-2 mutants had longer 3'-telomeric overhangs relative to wild type. These findings suggest that
clk-2 might be involved in telomere length regulation, and particularly may encode a single strand telomere binding protein. To explore this idea, we examined the C. elegans genome database for ORF's encoding potential regulators of telomere length. From our database search, we found that the ORF C07H6.6 shares some similarity with single strand telomere binding proteins in budding yeast, such as EST1, Cdc13p and TEL2. To determine whether C07H6.6 is the
clk-2 gene, we prepared a genomic region spanning the C07H6.6 by PCR and injected the fragment into
clk-2 (
qm37 ) animals. We found that C07H6.6 rescues
clk-2 mutant phenotype. We also used RNAi to confirm that C07H6.6 inhibition produces a phenotype similar to the
clk-2 mutation. From both DNA transformation rescue and RNAi, we conclude that the ORF C07H6.6 is the
clk-2 gene. To characterize the CLK-2 protein, we expressed the
clk-2 cDNA in Drosophila S2 cells. Currently, we are studying the biochemistry of CLK-2, and refining the phenotypes of
clk-2 mutants via adult life span, DNA damage response, apoptosis, and telomere clustering measurements. We will present our findings at the meeting.