The addition of ubiquitin (Ub) to a protein substrate (ubiquitination) is a critical regulatory post-translational modification, which can lead to degradation via the proteasome, translocation, or alteration of the activity of the targeted protein. C. elegans ARI-1 is a highly conserved member of the Ariadne (ARI) RBR E3 Ub-ligase subfamily. Though
ari-1 (C27A12.8) is expressed in numerous tissues including neurons, muscles, and the germline, strains homozygous for a null mutation in
ari-1(
tm2549) are viable and do not lead to obvious defects. The absence of a strong phenotype in
ari-1 mutants is likely due to genetic redundancy, as C. elegans possess two additional ari family members adjacent to
ari-1 (C27A12.6 and C27A12.7). Recently, our lab has utilized CRISPR technology to generate a null triple mutant in all three ari homologs [
ari3X]. Three independent strains were obtained that contain large deletions in the coding regions of C27A12.6, 7, and 8. Initial observations indicate a reduced brood size and ~25-60 % sterility in the
ari3X mutants. Sterile
ari3X mutants were found to have a normal soma but exhibit excessive sperm production, consistent with a Mog (masculinization of the germline) phenotype. Likewise, a null allele of
ubc-18(
tm5426), which encodes a conserved E2 partner of ARI proteins, also exhibits partial sterility and Mog. Furthermore, an independent screen recently reported a synthetic lethal interaction between
ubc-18 and fbf family members. Consistent with this, we find that both
ubc-18(
tm5426) and
ari3X mutants are hypersensitive to fbf(RNAi), resulting in a highly penetrant Mog phenotype. In addition,
ari3X; fbf(RNAi) worms exhibited a multivulval (Muv) phenotype and reduced germline proliferation (Glp),which have previously been observed in strong loss-of-function mutations in the fbfs. FBF-1 and FBF-2 are closely related RNA-binding proteins, which have the ability to bind thousands of different mRNAs and have been shown to regulate targets involved in germline development. Our data indicate that ARI-mediated ubiquitination plays an integral role in germline development and that ARI-UBC-18 functions coordinately with the FBFs to promote germ cell proliferation and oocyte differentiation. Studies are ongoing to determine whether ARI-UBC-18 affects FBF levels or co-represses specific germline targets with the FBFs that are involved in spermatogenesis.