ASNA1 is a highly conserved ATPase involved in a seemingly wide range of functions including metal resistance, growth control, insulin secretion and the targeting of tail-anchored proteins to membranes. In C. elegans, depleting ASNA-1 results in reduced insulin/IGF signalling (IIS) and in a growth arrest at the L1 stage without any apparent feeding defects. We wished to identify novel interactors to ASNA-1 that may shed light on the various roles of ASNA-1. Genetic studies, global yeast two-hybrid studies and co-immunoprecipitation studies have led to the identification of proteins that interact with ASNA1 homologues in various model systems. We have undertaken a feeding RNAi screen on the homologues of these genes in C. elegans to identify genes that, like
asna-1, cause a larval growth arrest in stage L1 with reduced IlS upon depletion by RNAi. Using bioinformatics we assembled a list of 143 possible ASNA-1-interactors, based on interaction maps from flies, worms and yeast. By means of feeding RNAi against 98 of these genes, we were able to identify six that upon inactivation caused
asna-1-like L1 arrests:
rps-0 (ribosomal SA subunit B0393.1), T23B12.3 (mitochondrial ribosome S2 subunit)
iars-1 (iso-leucine aminoacyl tRNA synthetase),
ykt-6 (synaptobrevin), F41C3.4 (homolog of the human vesicle transport gene Got1), and
goa-1 (G protein alpha subunit Go). RNAi against two of those genes,
rps-0 and T23B12.3 led to decreased insulin signaling (using the DAF-16:GFP localization assay) in addition to the larval arrest. RNAi against T23B12.3 enhances synthetic semi-dauer formation in
daf-7(
e1372) mutants and enhances the L1 arrest phenotype of
asna-1 mutants in a sensitized background.
iars-1(rnai) produces a strong L1 arrest without any feeding defects, but we are unable so far to find any evidence for an insulin signaling defect in the worms depleted for the gene activity.
ykt-6(rnai) animals show defects in insulin signaling but also display moderate feeding defects as well. RNAi against
goa-1 and the GOT1 homolog led to severe feeding defects and were not considered further. Both
rps-0 and T23B12.3 are evolutionarily well conserved, raising the possibility that our screen may identify candidates that affect insulin signaling in vertebrates.