The biochemical nature of the hatching substance released by Zlyanassa obsoleta embryos was examined by assessing the biological activity of the hatching substance after several treatments. Active preparations of the hatching substance capable of dissolving the egg capsule plug were inactivated in sodium dodecyl sulfate, 2-mercaptoethanol, some proteases, EGTA, and upon boiling. Coincident with the release of the hatching substance from embryos was the appearance of several proteins and protease activity. However, specific protease inhibitors did not abolish hatching activity. The hatching substance had maximum activity at pH 7.0 and 20°C and may have a native molecular weight in excess of 100,000 daltons. These characteristics of the Zlyanassa hatching substance are very similar to the properties of the well-studied hatching enzymes of several other embryos, yet it is still uncertain if the hatching substance is functioning as an enzyme.Extracellular envelopes surround the developing embryos of most animals. These envelopes range from relatively simple coverings such as the acellular zona pellucida of mammalian embryos to the elaborately patterned chorion of fish embryos. Embryonic development proceeds until the time of hatching, when the embryo emerges from the envelope. Although hatching usually has a mechanical component (various movements of the embryo's body), the process is often chemically mediated. Proteolytic enzymes are responsible for the emergence of sea urchin blastulae from the fertilization envelope (Barrett and Edwards, '76; Edwards et al., '77; Hoshi et al., '79), fish embryos from their chorion (Yamagami, '73; '81; DiMichele et al., '81; Schoots and Denuce, ' €311, and amphibian embryos from the fertilization envelope (Katagiri, '75; Urch and Hedrick, '81). Thus far, most of the research on hatching has been restricted to studies of systems in which embryos are hatching from envelopes produced in the ovary (primary envelopes produced by the egg itself or secondary envelopes produced by surrounding follicle cells) Wilson, '25). Chemical hatching from envelopes secreted by cells of the oviduct (tertiary envelopes) has yet to be studied in such detail.One example of a tertiary envelope is the egg capsule of many gastropod molluscs. Egg capsules are often large structures (from 1 mm to several cm in length), containing from one to several thousand embryos. Along with osmotic and mechanical hatching mechanisms, chemical hatching has been described for gastropods (reviewed by Davis, '68). During chemically mediated hatching, a predesignated region of the capsule, the operculum or plug, dissolves, producing an opening through which the embryos escape to take up a larval or juvenile life. Hatching enzymes have been suggested, though not clearly demonstrated, as being responsible for hatching in Urosalpinx cinerea (Hancock, '56), Cerithium algicola (Davis, '67), Zlyanassa obsoleta (Pechenik, '751, and Busycon carica and B. canaliculatum (Harasewych, '78).Because Zlyanassa is readily adapt...
