Pupal cuticle proteins of Manduca sexta: characterization and profiles during sclerotization

Proteins in pupal abdominal cuticle of the tobacco hornworm, Manduca sexta, were characterized during the pre-ecdysial and post-ecdysial periods of sc lerotization and endocuticle formation. Protein extractability decreased dr amatically as the cuticle became sclerotized through 6 h post-ecdysis, but increased rapidly from 9 to 48 h as endocuticular layers were secreted. Nea rly 100 proteins that were extracted from pre-ecdysial cuticle became large ly insoluble during sclerotization. Three major proteins in this group dest ined to become exocuticle had apparent molecular masses (Mapp) of 20, 27 an d 36 kDa , and were designated MS-PCP20, MS-PCP27, and MS-PCP36, Amino acid analysis revealed glycine to predominate in all three proteins, and alanin e, aspartate, glutamate, proline and serine were also relatively abundant. Histidine residues, which provide sites for adduct and cross-link formation with quinone metabolites of N-beta-alanyldopamine during sclerotization of pupal cuticle, ranged from 2 to 3 mol %. N-Tenninal amino acid analysis of MSPC-20 and MSPC-36 also revealed some sequence similarities indicating th ey may be related. An almost entirely new group of proteins appeared by 9 h as endocuticule secretion began, and these increased in abundance through 48 h post-ecdysis. Two of these were major proteins with Mapps of 33 and 34 kDa, and they also had close similarities in their N-terminal amino acid s equences. This study showed that the large number of proteins secreted into the presumptive exocuticle of the pupa before ecdysis are involved in scle rotization reactions and as a consequence become largly insoluble. The epid ermis then switches to the secretion of an entirely new group of proteins t hat are involved in formation of the endocuticle. (C) 2000 Elsevier Science Ltd. All rights reserved.