Pea Crabs and Barnacles: Egg Hatching, Peptide Pheromones, and Endoproteinases

Abstract

Barnacles, pea crabs and other crustaceans brood eggs. Brooding enables females to manage biofouling on the eggs and ventilate them. Though glued to appendages on the abdomen, there is no physiological connection between embryos in eggs and the brooding female. Females use enzyme products as pheromones to coordinate behavioral, physical, and biochemical processes involved in egg hatching and larval release. Larval release pheromones are peptides generated by exogenous trypsins acting on proteins. Peptide pheromones coordinate egg hatching and larval release, which is synchronized between the embryos and the female. Eggs are brooded during embryo development and are physically and enzymatically cleaned until egg hatching. My interest was to identify the endoproteinases associated with pea crabs and barnacles. Peptides generated from hydrolysis of pure proteins were identified by high resolution LC electrospray, MS, MS. Using pure proteins enables absolute identity of the sequences around the proteolytic cleavages, which allows for definition of enzyme specificity. Proteomics analysis identified peptides from pure proteins generated by maintenance proteases and proteases involved in egg hatching and larval release. The enzymatic activity in the host plus microbiome communities of pea crabs and barnacles is much greater and has a different pattern than enzymes in seawater. Enzyme patterns of the functional microbiomes are similar within each species and different between species. The addition of proteins to pea crabs with late stage embryos and barnacles with late stage embryos triggered hatching. Based on carboxyl terminus amino acids of peptides, there is an ensemble of 14 endoproteinases with major contributions from serine proteases.