Elucidating the mechanisms of blister localization in bli-5 mutant Caenorhabditis elegans

In C. elegans, the gene bli-5 encodes a serine-protease inhibitor that is responsible for blocking the proteases nas-36 and nas-37, both of which are involved in ecdysis and cuticle assembly during molting. Mutations in bli-5 result in the formation of fluid-filled blisters along the cuticle of the worm. In contrast to mutations in the 5 other genes that result in the blistered phenotype, blisters caused by defects in bli-5 form only on the head of the worm. Transcriptional analysis of bli-5 has shown that it is expressed at high levels in a singular cell within the C. elegans excretory system. Fluorescence microscopy demonstrated that this is the gland cell, which is a binucleate syncytium responsible for production and secretion of unknown intracellular molecules. Secretions from the gland cell have been implicated in molting in the nematode Phocanema decipiens, corroborating the findings that bli-5 would be produced and secreted from this cell. In a separate experiment, RNAi bacteria and mutant worms were used to create worms deficient in both nas-36 and nas-37. Single-mutants in either of these genes results in a cuticle-dragging phenotype, indicating that the two genes do not have entirely redundant functions. In order to better understand the individual functions of nas-36 and nas-37, we examined the aforementioned double-knockout worms to assess viability and molting function, determining that molting function was still preserved, although the growth of the worm was delayed significantly. This implicates both nas-36 and nas-37 as having important roles in shedding the cuticle, but more importantly indicates that they are not the only proteases involved in removing old cuticle during ecdysis, as molting still occurs in the absence of both genes. To better understand the molting process, characterization of the other genes involved in ecdysis and their regulators would be immensely beneficial