In Caenorhabditis elegans , locomotion is driven by the transmission of force generated by body wall muscles across basal lamina and hypodermis to the cuticle. The mechanical pathway, whereby skeletal muscles transmit contractile force, is composed of various cytoskeletal proteins and matrix components. Mutations that affect normal assembly or integrity of this linkage system exhibit a paralyzed phenotype. Mutated genes of the mua ( m uscle a ttachment) class result in a progressive paralysis due to post-embryonic failure of the structural linkages during normal use. To date, all mua -class genes appear to affect hypodermal links within this pathway. Specific mutations result in detachment initiated at characteristic locations–adhesion of muscle to hypodermis, adhesion of hypodermis to cuticle, or integrity of the hypodermis. One mua gene of interest is the
vab-10 gene. Mutations in this gene result in viable worms that are only partially paralyzed, and are characterized by a "bent head" phentotype. In
vab-10 mutants, muscle assembly and differentiation are normal, but are followed by spontaneous muscle detachment from the body during growth.
vab-10 maps to an interval of chromosome I that contains the C. elegans plectin and kakapo homologs. Both of these genes are contained within the cosmid ZK1151. We injected this cosmid into animals and found that ZK1151 rescues
vab-10 mutants. The ZK1151 cosmid houses three predicted genes–ZK1151.1 encodes an unknown protein with spectrin-like repeats, while ZK1151.2 and ZK1151.3 encode for proteins with kakapo and plectin--like homology. Both the plectin and kakapo-like genes prove to be promising candidates for
vab-10 . Using restriction digestion followed by religation, specific genes are being knocked out of the ZK1151 cosmid and the resulting subclones microinjected into mutant
vab-10 animals to assess rescue and the identity of the
vab-10 gene. Further characterization of
vab-10 will also include immunological techniques. The primary site of muscle detachment in
vab-10 mutants is believed to be between hypodermis and muscle. Recent work with GFP-tagged intermediate filaments has shown that
vab-10 mutants exhibit a pulling away of the muscle from the body wall with the hypodermis remaining attached to the cuticle (Wura Omotosho, personal communication). Whether peeling occurs from muscle and basal lamina or from hypodermis and basal lamina, and if other structures are affected will be determined with further immunostaining. In addition, other mua genes will be examined using immunofluorescent probes for specific muscle, basal lamina, and hypodermal proteins including
mua-1 ( encoding a transcription factor) and
mua-3 (encoding a protein associated with fibrous organelles).