Uncoupling the Pleiotropic Phenotypes of clk-1 with tRNA Missense Suppressors in Caenorhabditis elegans

Abstract

clk-1 encodes a demethoxyubiquinone (DMQ) hydroxylase that is necessary for ubiquinone biosynthesis. When Caenorhabditis elegans clk-1 mutants are grown on bacteria that synthesize ubiquinone (UQ), they are viable but have a pleiotropic phenotype that includes slowed development, behaviors, and aging. However, when grown on UQ-deficient bacteria, the mutants arrest development transiently before growing up to become sterile adults. We identified nine suppressors of the missense mutation clk-1(e2519), which harbors a Glu-to-Lys substitution. All suppress the mutant phenotypes on both UQ-replete and UQ-deficient bacteria. However, each mutant suppresses a different subset of phenotypes, indicating that most phenotypes can be uncoupled from each other. In addition, all suppressors restore the ability to synthesize exceedingly small amounts of UQ, although they still accumulate the precursor DMQ, suggesting that the presence of DMQ is not responsible for the Clk-1 phenotypes. We cloned six of the suppressors, and all encode tRNA^Glu genes whose anticodons are altered to read the substituted Lys codon of clk-1(e2519). To our knowledge, these suppressors represent the first missense suppressors identified in any metazoan. The pattern of suppression we observe suggests that the individual members of the tRNA^Glu family are expressed in different tissues and at different levels.

We thank David Shanks for his work in the early stages of the project, particularly for helping to isolate mutants. We gratefully acknowledge C. F. Clarke, P. N. Rather, and M. Kawamukai for kindly providing bacterial strains and the Caenorhabditis Genetics Centre, which is funded by the National Institute of Health National Center for Research Resources (NIH NCRR), for providing nematode strains.

This work was funded in part by a research contract from Chronogen Inc. R.B. was supported by a Mary Louise Taylor McGill Major Fellowship, P.A.T.N. by a McGill Faculty of Graduate Studies Fellowship, and S.H. is a Strathcona professor of zoology.