Cloning and Expressional Analysis of FKBP12 in Crustacean Skeletal Muscle

Molting in decapod crustaceans is stimulated by ecdysteroids secreted by the Y-organs. During premolt, the claw muscle must atrophy to successfully withdraw the claws from the exoskeleton. This study investigates the cellular mechanisms responsible for this muscle atrophy. In the blackback land crab, Gecarcinus lateralis, the rate of global protein synthesis in atrophic claw muscles is correlated with hemolymph ecdysteroid titers. The increased protein turnover is associated with extensive remodeling of the contractile structure as muscle fibers are reduced in size. These data suggest that ecdysteroids stimulate metazoan Target Of Rapamycin (mTOR), a protein kinase complex that promotes translation. We hypothesize that FKBP12 (FK506-binding protein, 12 kDa), a known inhibitor of mTOR, is down regulated in atrophic claw muscle. FKBP12 from European green crab (Carcinus maenas) was sequenced and the aligned with other Crustacean species and across phyla. qPCR was used to quantify the effects of eyestalk ablation and multiple leg autotomy on FKBP12 expression in claw and thoracic muscles of C. maenas. The animals were refractory to molt induction, and remained in intermolt the entire duration of each experiment. There was no consistent pattern of FKBP12 expression. In C. maenas that molted naturally, there was an aggregated 3-fold increase of FKBP12 expression in claw muscle as compared to thoracic muscle. The naturally molting animals had a 100-1,000 fold decrease in FKBP12 expression in both claw and thoracic muscle compared to the non-molting animals. Higher FKBP12 expression in the refractory animals could implicate a place for FKBP12 in controlling molt suspension.