The lifespan of the nematode Caenorhabditis elegans is genetically determined by several classes of genes. Our laboratory is particularly interested in the clk class of genes, which is composed of
clk-1,
clk-2,
clk-3, and
gro-1. Mutations in these genes have been shown to extend lifespan and to deregulate several developmental and behavioral processes. For example,
clk-1 mutants exhibit an average slowing down of cell division, pharyngeal pumping, and defecation rates relative to wild-type animals. These phenotypes are maternally rescued, that is homozygous mutant animals coming from a heterozygous mother are phenotypically wild type.
clk-1 encodes a 187 amino acid mitochondrial protein that is composed of two homologous TRC domains (TRC for Tandemly Repeated in CLK-1). Interestingly, the yeast homologue of
clk-1, COQ7, has been implicated in ubiquinone biosynthesis. Moreover,
coq7 mutants are respiration defective, yet the respiratory competence can be restored by the addition of exogenous ubiquinone. In contrast, cellular respiration is only slightly affected in
clk-1 mutant worms. However, C. elegans
clk-1 as well as the rat and the human homologues are capable of functionally complementing the Dcoq7 mutant. This observation suggests that the biochemical function of
clk-1 have been conserved throughout evolution. We have shown previously that a recombinant CLK-1::GFP fusion protein is capable of rescuing the
clk-1 mutant phenotype when expressed from transgenic arrays. In the current study, we are taking advantage of this observation to try to understand the structural requirements for CLK-1 function. In order to address this, we are using a site-directed mutagenesis approach to generate several new CLK-1 mutant proteins. We are analysing the effect these CLK-1 mutant proteins have on the general
clk-1 phenotype. We are also examining the subcellular distribution of the mutant fusion proteins in different genetic backgrounds, such as the wild type, the
clk-1 partial loss of function
e2519, and the two
clk-1 putative nulls
qm30 and
qm51.