ZHOU, J.-B.; MEDLER, S.; CHANG, E. S.; MYKLES, D. L.; Colorado State University; Colorado State University; Bodega Marine Laboratory; Colorado State University: Cloning and Expression of Lobster Calcineurin A

Calcineurin is a Ca2+-activated protein phosphatase involved in Ca2+-dependent signal transduction in various tissues; known targets are histones, myosin light chain, cAMP-dependent protein kinase, and transcription factors. In mammalian skeletal muscle it mediates the induction of slow fiber genes in response to chronic motor neuron activity. The purpose of this study is to determine the role of calcineurin in fiber-type-switching in the American lobster, Homarus americanus. Fibers in the claw closer muscles undergo a developmentally-regulated switching as the isomorphic claws of larvae and juveniles differentiate into the heteromorphic cutter and crusher claws of the adult. A cDNA encoding a partial calcineurin A sequence was obtained with RT-PCR using degenerate primers to a highly-conserved sequence in homologous genes from other species. The expression of calcineurin in different lobster tissues was examined by RT-PCR, real time PCR and Northern blot analysis. The RT-PCR product (449 bp) was sequenced to confirm that it was calcineurin A, and the sequence was used to design sequence-specific primers for RT-PCR and real time PCR. RT-PCR indicated that calcineurin was expressed in all tissues examined, including skeletal muscle, heart, digestive gland, testis and intestine. Real time PCR showed that there was no difference in the levels of expression between slow fibers in crusher claw muscle and fast fibers in cutter claw and deep abdominal muscle from intermolt animals. Preliminary results from Northern blot analysis showed that the size of the calcineurin A transcript was about 2.7 kb. Future studies will examine expression claw muscles during development and during the molting cycle. Supported by NSF (IBN 00-77422).