Distribution of serpins in the tissues of the tobacco hornworm (Manduca sexta) larvae - Existence of new serpins possibly encoded by a gene distinct from the serpin-1 gene

Polyclonal antibodies were raised against the isolated hemolymph serine pro teinase inhibitors (serpins) of Manduca sexta larvae. Two of these antibodi es, MsH49a and MsH49b, displayed characteristic differences in labelling pa tterns of hemocytes, fat body, integumental epidermis and cuticle on immuno blots, and in light- and electronmicroscopic sections. The serpin compositi on of the latter three tissue homogenates was determined by native immunobl ots and inhibitor binding assays. The results were compared to the hemolymp h samples containing all the known inhibitors encoded by the well-character ized serpin-1 gene. The enzyme specificity of the MsH49b-labelled cuticular serpin was similar to serpin-1J, although its electrophoretic mobility on native PAGE was not identical with any of the known proteinase inhibitors e ncoded by the serpin-1 gene. Based on these data, we suggest that the cutic le and hemolymph may contain novel serpin(s) encoded by a gene other than t he serpin-1 gene. Since the serpin-1J proved to be involved in the activati on pathway of the prophenoloxidase system in the hemolymph, the in vivo fun ction of cuticular MsH49b serpin was investigated by prophenoloxidase tests in native cuticular homogenates. Our results demonstrated that the cuticul ar serpin(s) that are labelled by the MsH49b antibody may play a determinan t role in the regulation of the prophenoloxidase system of the integumental cuticle. (C) 2001 Elsevier Science Ltd. All rights reserved.