WHEATLY, M.G.*; GAO, Y.: Crayfish Plasma Membrane Ca ATPase: Characterization, Expression and Evolution.

The crayfish molting cycle is used to study regulation of expression for Plasma Membrane Ca ATPase (PMCA). As in vertebrates PMCA is encoded by a family of 4 genes (PMCA 1-4). Expression was examined in an epithelium that transports Ca (gill) versus a control tissue (axial muscle). PMCA1 exhibited one “housekeeping” isoform that was expressed uniformly during the molting cycle. Two PMCA2 isoforms were amplified in each tissue. PMCA2b was expressed throughout all stages; however PMCA2a was upregulated in pre- and postmolt. PMCA3 also exhibited two isoforms, both of which were upregulated in pre- and postmolt. PMCA4 showed different expression patterns in gill versus muscle. In muscle a single isoform (PMCA4b) was observed. In gill PMCA4b was upregulated in premolt and PMCA 4b in postmolt. Within the PMCA family of genes some interesting patterns are emerging that may correlate with enhanced transepithelial Ca transport during postmolt. Two PMCA genes have been partially cloned and sequenced (PMCA1, 2500 bp; PMCA2, 2764 bp). A polyclonal antibody against PMCA2 recognized a prominent band at 110 kDa in Western blots of membranes isolated from egg, gill, antennal gland, hepatopancreas and muscle. Immunocytochemistry confirmed antibody hybridization to basolateral membranes in antennal gland. A phylogenetic analysis of crayfish partial PMCAs using the Jotun Hein method of MEMALIGN indicated that crayfish PMCAs exhibit common evolutionary relationships with the vertebrate PMCA gene family. (Supported by NSF grant IBN 0076035).