IBN 98-08156 DONG The goals of this research are 1) to study the molecular basis of interactions between the German cockroach sodium channel (the protein molecule that forms pores in cell membranes which selectively allow sodium ions to enter and leave the cell) and pyrethroid insecticides, and 2) to examine the functional properties and developmental and tissue-specific expression (that is, which tissues have these) of alternatively spliced isoforms of the German cockroach para gene (alternatively spliced isoforms are different versions of a protein, just the way a car manufacturer may have different models within a particular line of cars). Pyrethroid insecticides affect the properties of voltage-dependent sodium channels in the nervous system (that is the sodium channels that are responsible for sustaining the nerve impulses through which nerve cells communicate). A class of so-called kdr (knockdown resistance) mutations has been identified in several insects. kdr insects are resistant to knockdown (rapid paralysis) and kill by pyrethroids and some sodium-channel site-2 neurotoxins. Recently para genes from kdr and genetically related pyrethroid-susceptible German cockroach strains have been cloned and sequenced (that is, their genetic code has been deciphered). Sequence comparison reveals specific amino acid differences between the Para proteins from kdr and those from susceptible cockroach strains. The first objective of this research is to compare the sensitivities of wild-type (susceptible) and mutant Para sodium channels to pyrethroids and site-2 neurotoxins to confirm that the putative kdr mutations are indeed responsible for cockroach knockdown resistance to pyrethroids. "Alternative splicing" (in which certain parts of a molecule can be selected from various options, much as parts of a car such as the engine or the radio might be taken from a set of possible options) is a common feature of sodium channel genes of both vertebrates and invertebra tes. The functional significance of alternatively spliced variants, however, remains largely unknown. Seven alternatively spliced portions of the cockroach para gene have been identified, three of which are unique among insects. The second objective of this research is to examine the functional properties of unique para splice variants and to analyze their developmental and tissue-specific expression patterns. Results from this project will enable us to understand why insects become resistant to insecticides as well as the biological significance of alternatively spliced Para channel variants in governing sodium channel properties.
