PROTEINS OF CRUSTACEAN EXOSKELETON .2. IMMUNOLOGICAL EVIDENCE FOR THEIR RELATEDNESS TO CUTICULAR PROTEINS OF 2 INSECTS

The exoskeletons of crustaceans and cuticles of insects are characterized by an abundance of proteins with acidic pIs and Ms 31 kDa or smaller. Immunochemical data described here further demonstrate the similarities of such proteins among representatives of these two major classes of Arthropoda. Proteins extracted from the four individual layers of the exoskeletons of Bermuda land crabs Gecarcinus lateralis at different stages of the intermolt cycle and from exuviae, the partially degraded old exoskeleton that is cast at ecdysis, were analyzed on Western blots with polyspecific polyclonal antibodies against either larval cuticular proteins (anti-LCP) or pupal cuticular proteins (anti-PCP) of the tobacco hornworm Manduca sexta, or against two pairs of larval cuticular proteins from third instar larvae (anti-L3CPS) of Drosophila melanogaster. M. sexta anti-LCP cross-reacted with many of the crab proteins extracted at all stages of the intermolt cycle. It reacted with more high M(r) proteins from late proecdysial exoskeleton and exuviae than with such proteins from anecdysial or early proecdysial exoskeletons, probably because the larger M, proteins are enriched when the lower M(r) proteins are degraded during proecdysis. M. sexta anti-PCP cross-reacted hardly at all with the crab proteins. Both D. melanogaster anti-L3CPS cross-reacted with crab epicuticular proteins at all stages of the intermolt cycle and from exuviae. Anti-L3CP 3 + 4 reacted even more intensely and with many more proteins from the three inner layers of the exoskeleton than did anti-L3CP 1 + 2. Controls included 2D gels with rabbit anti-ovalbumin to detect glycoproteins containing mannose and N-acetylglucosamine; it cross-reacted with only one protein in membranous layer of an anecdysial crab, while rabbit anti-fluorescein reacted with none. By contrast, the intensity and specificity of the cross-reactions of crab exoskeletal proteins with antibodies against cuticular proteins of members of two orders of insects suggest that at least some cuticular proteins in the two arthropod Classes, Crustacea and Insecta, are evolutionarily conserved.