We propose to analyze the assembly of the spermatophore, the sac that packages the sperm for transfer from male to female in the mealworm beetle. In this beetle, as in many other arthropods, secretions from the male reproductive accessory glands flow into the ejaculatory duct where they are molded into the multilayered spermatophore. Because much is known about its accessory glands, the mealworm beetle is an especially favorable model with which to study accessory gland development and spermatophore formation. The formation of a spermatophore requires multi-step aggregation of monomers into aggregates that ultimately are assembled into networks. First, we will characterize representative structural proteins, termed spermatophorins, that are produced by the accessory glands. With the aid of a monoclonal antibody, we have isolated one spermatophorin, in a proline-rich protein of 23 kD designated Sp23, and we have begun to characterize it and to determine its amino acid sequence by Edman degradation and by inference from the corresponding cDNAs. From sequence information and physical data, we can inter some of the structural characteristics of individual spermatophorins. With antibodies, we can determine which of these co-localize in the spermatophore. To investigate the formation of homo-and heteroaggregates, we will use symmetrial bifunctional cross- linkers. To identify the ligands which bind to a given spermatophorin (such as Sp23), we will employ heterobifunctional cross-linkers. We hope to investigate the kinetics of their interactions, and to define the domains which link these proteins to one another. Our conclusions about mechanisms of assembly may be relevant to analogous assembly processes in cuticle and chorion. In addition, we hope to collaborate with others in the application of our antibodies and techniques to spermatophores of other genera, including such as Simulium, Culicoides, and Glossina, the vectors of filaria, blue-tongue virus, and trypanosomes, respectively. Increased knowledge of the reproductive biology of these vectors may aid in the development and evaluation of pest control programs.
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