Post-translational modification of proteins by ubiquitination is essential for viability and development in eukaryotic organisms. Ubiquitin, a highly conserved 76 amino acid peptide, is covalently conjugated to various target proteins at lysine residues through the sequential action of an E1 ubiquitin-activating enzyme, an E2 ubiquitin-conjugating enzyme, and an E3 ubiquitin ligase. Poly-ubiquitination typically results in target protein degradation via the proteasome. We report here the cloning and characterization of a temperature-sensitive allele of the E1 activating enzyme encoded by
uba-1 in C. elegans. The allele
it129 (previously known as
spe-32) was mapped by three-factor crosses and its position further refined by snip-SNPs. Candidate genes within the interval were screened by RNAi feeding. A single construct corresponding to
uba-1 reproduced multiple phenotypes of the
it129 mutation. A deletion allele of
uba-1,
ok1374, failed to complement
it129. PCR amplification and sequence determination of the
uba-1 coding region from
it129 revealed a single missense mutation that, based on structural data, maps near the ubiquitin binding site of the E1 protein. Homozygous mutants of
uba-1(
it129) manifest different phenotypes depending upon the timing of the temperature shift, and include embryonic lethality, larval lethality, and sperm-specific sterility. Males exhibit additional sex-specific phenotypes, with malformation of mating structures in the tail and progressive paralysis culminating in premature death. The latter phenotype is particularly intriguing as it occurs post-developmentally, and may provide an attractive system to study age-related declines in neuromuscular function. The
uba-1 gene is expressed in a variety of cells throughout development, consistent with the multiple phenotypes observed. We propose a model whereby proteins normally targeted for proteasomal degradation instead persist in
uba-1(
it129) and impair critical cellular processes as a consequence.