KIM, Hyun Woo; YU, Xiaoli ; MYKLES, donald L; Colorado State University: Characterization of cDNAs encoding decapod crustacean calpains

Calpains constitute a large and diverse family of Ca2+-dependent cysteine proteinases in animal cells. In decapod crustaceans, four calpain activities mediate molt-induced claw muscle atrophy. Selective degradation of myofibrillar proteins reduces muscle mass so that the claws are easily withdrawn during ecdysis. We previously characterized a muscle-specific calpain (Ha-CalpM) in lobster (Yu , X.-L. and Mykles, D.L. , Am. Zool. 41: 1634; 2001). We have now isolated additional calpain species using RT-PCR and 3?RACE. cDNAs encoding three calpain genes (Gl-CalpA, Gl-CalpB, and Gl-Calp5) from land crab (Gecarcinus lateralis) limb regenerates and one from American lobster (Homarus americanus) skeletal muscle (Ha-CalpB) were characterized. Gl-CalpA had high homology with the Ha-CalpM sequence and was expressed as two alternatively-spliced transcripts. The CalpB genes from lobster and crab have a calmodulin-like domain at the C-terminus. Gl-Calp5 sequence was similar to human type 5 calpain and nematode TRA-3. Results from RT-PCR and Northern blot analysis showed that Gl-CalpA is predominantly expressed in skeletal muscles. CalpB and Gl-Calp5 had a ubiquitous tissue distribution, with Gl-Calp5 expressed at high levels in X organ. Real time PCR showed that calpain mRNA levels change over the molting cycle. Supported by NSF (IBN-9904528).