During the juvenile-to-adult transition at the L4 stage, the tail tip cells in C. elegans males fuse, change shape and migrate anteriorly. The timing of this tail tip morphogenesis (TTM) is governed by the heterochronic pathway, best known for scheduling seam cell development. Previous work showed that heterochronic genes
let-7 and
lin-41 play a role in the timing of TTM, which is delayed into adulthood in
let-7(lf) and
lin-41(gf) mutants. We identified two new heterochronic pathway genes,
lep-2 and
lep-5. Males mutant in either gene have fully unretracted ("leptoderan", Lep) tail tips that delay TTM into adulthood. Additionally, mutant adults undergo an additional molt and mutant
lep-2 males show defective mating behavior, suggesting that the genes act in the body epidermis and nervous system as well as the male tail. Neither mutant has obvious defects in lateral seam development.
lep-2 encodes a Makorin, a member of a family of conserved proteins with putative nucleic acid binding and E3 ubiquitin ligase activities. The active product of
lep-5 is a cytoplasmically localized, long noncoding RNA (lncRNA), well conserved within Caenorhabditis but not yet identified outside of this genus. SL1-spliced
lep-5 lncRNA is predicted to adopt a secondary structure with several stem-loops, at least two of which are necessary for its function. Using a photo-convertible LIN-28::Dendra2 fusion, we found that LEP-2 and
lep-5 lncRNA promote degradation of LIN-28 protein, without affecting
lin-28 mRNA levels. This is consistent with the predicted E3 ubiquitin ligase activity of LEP-2, which also has a predicted RNA-binding domain. Specifically, we hypothesize that the
lep-5 lncRNA serves as a scaffold to bring LEP-2 and LIN-28 into close proximity to allow ubiquitination of LIN-28, tagging it for proteasomal degradation. We are currently testing this hypothesis. The proposed mechanism could be conserved in vertebrates. The human homolog of LEP-2, MKRN3, and human LIN28B, are both involved in the regulation of pubertal timing. Together with our findings in C. elegans, this suggests a conserved role of Makorins and LIN-28 in timing of the juvenile-to-adult transition. If a mammalian homolog of
lep-5 lncRNA exists, it is expected to show structural similarity rather than sequence similarity; we are searching for such structural homologs. The fact that seam cell development is apparently unaffected by mutations in
lep-2 and
lep-5 suggests that developmental timing is regulated in a tissue-specific manner. Therefore, the male tail tip is a valuable model for studying the heterochronic pathway in all its complexity.