Abstract
The conserved Caenorhabditis elegans protein kinases NEKL-2 and NEKL-3 regulate membrane trafficking and are required for larval molting. Through a forward genetic screen we identified a mutation in catp-1 as a suppressor of molting defects in synthetically lethal nekl-2; nekl-3 double mutants. catp-1 encodes a membrane-associated P4-type ATPase involved in Na+-K+ exchange. A previous study found that wild-type worms exposed to the nicotinic agonist dimethylphenylpiperazinium (DMPP) exhibited larval arrest and molting-associated defects, which were suppressed by inhibition of catp-1. By testing a spectrum catp-1 alleles, we found that resistance to DMPP toxicity and the suppression of nekl defects did not strongly correlate, suggesting key differences in the mechanism of catp-1-mediated suppression. Through whole genome sequencing of additional nekl-2; nekl-3 suppressor strains, we identified two additional coding-altering mutations in catp-1. However, neither mutation, when introduced into nekl-2; nekl-3 mutants using CRISPR, was sufficient to elicit robust suppression of molting defects, suggesting the involvement of other loci. Endogenously tagged CATP-1 was primarily expressed in epidermal cells within punctate structures located near the apical plasma membrane, consistent with a role in regulating cellular processes within the epidermis. Based on previous studies, we tested the hypothesis that catp-1 inhibition induces entry into the pre-dauer L2d stage, potentially accounting for the ability of catp-1 mutants to suppress nekl molting defects. However, we found no evidence that loss of catp-1 leads to entry into L2d. As such, loss of catp-1 may suppress nekl-associated and DMPP-induced defects by altering electrochemical gradients within membrane-bound compartments.