Caenorhabditis elegans molting is a process during which the apical extracellular matrix of the epidermis, the cuticle, is remodeled through a process of degradation and re-synthesis. Using a genetic approach, we identified nekl-3 as essential for the completion of molting. NEKL-3 is highly similar to the mammalian NEK kinase family members NEK6 and NEK7. Animals homozygous for a hypomorphic mutation in nekl-3, sv3, had a novel molting defect in which the central body region, but not the head or tail, was unable to shed the old cuticle. In contrast, a null mutation in nekl-3, gk506, led to complete enclosure within the old cuticle. nekl-2, which is most similar to mammalian NEK8, was also essential for molting. Mosaic analyses demonstrated that NEKL-2 and NEKL-3 were specifically required within the large epidermal syncytium, hyp7, to facilitate molting. Consistent with this, NEKL-2 and NEKL-3 were expressed at the apical surface of hyp7 where they localized to small spheres or tubular structures. Inhibition of nekl-2, but not nekl-3, led to the mislocalization of LRP-1/megalin, a cell surface receptor for low-density lipoprotein (LDL)-binding proteins. In addition, nekl-2 inhibition led to the mislocalization of several other endosome-associated proteins. Notably, LRP-1 acts within hyp7 to facilitate completion of molting, suggesting at least one mechanism by which NEKL-2 may influence molting. Notably, our studies failed to reveal a requirement for NEKL-2 or NEKL-3 in cell division, a function reported for several mammalian NEKs including NEK6 and NEK7. Our findings provide the first genetic and in vivo evidence for a role of NEK family members in endocytosis, which may be evolutionarily conserved.
Bibliographical noteFunding Information:
Cosmid clones were generously provided by Audrey Fraser, Ratna Shownkeen, and Alan Coulson of the Wellcome Trust Sanger Institute, Hinxton, UK. We thank Barth Grant for generously providing C. elegans strains. Some strains were provided by the CGC, which is funded by the US National Institutes of Health (NIH) Office of Research Infrastructure Programs ( P40 OD010440 ). We are also greatly indebted to Bob Herman and Simon Tuck for their generous support of this project, to Amy Fluet for editing and to Jay Gatlin for help with microscopy. Support at the University of Minnesota was from NIH (Grant GM22387 to LC). Support at the University of Wyoming was from NIH (Grant GM066868 to DSF).
© 2014 Elsevier Inc.
- C. elegans