Cysteine proteinases are involved in a variety of important biological processes and have been implicated in molting and tissue remodeling in free living and parasitic nematodes. We show that in the lymphatic filarial nematode Brugia pahangi molting of third-stage larvae (L3) to fourth-stage larvae is dependent on the activity of a cathepsin L-like cysteine protease (CPL), which can be detected in the excretory/secretory (ES) products of molting L3. Directed cloning of a cysteine protease gene in B. pahangi and analysis of the expressed sequence tag (EST) and genomic sequences of the closely related human lymphatic filarial nematode Brugia malayi have identified a family of CPLs. One group of these enzymes, Bm-cpl-1, -4, -5 and Bp-cpl-4, is highly expressed in the B. malayi and B. pahangi infective L3 larvae. Immunolocalization indicates that the corresponding enzymes are synthesized and stored in granules of the glandular esophagus of L3 and released during the molting process. Functional analysis of these genes in Brugia and closely related CPL genes identified in the filarial nematode Onchocerca volvulus and the free living model nematode Caenorhabditis elegans indicate that these genes are also involved in cuticle and eggshell remodeling.