SICB Division of Comparative Physiology and Biochemistry (DCPB)

Researchers Database Entry

Donald Mykles

Regulation of Molting and Skeletal Muscle Plasticity in Crabs and Lobsters
Our research concerns the regulation of molting and skeletal muscle plasticity in crabs and lobsters. Specific areas are signaling mechanisms in the molting gland and proteolytic mechanisms mediating molt-induced claw muscle atrophy. Quantitative methods are used to measure transcriptional and post-transcriptional regulation of signal transduction of molt-inhibiting hormone (MIH), a neuropeptide that inhibits ecdysteroid synthesis in the molting gland and myostatin, a TGFb superfamily member that induces skeletal muscle atrophy.

Signaling mechanisms in crustacean molting gland. Growth in crustaceans requires the periodic shedding of the shell, a process called molting, which is controlled by a neurosecretory center in the eyestalks. The complex secretes MIH that inhibits production of the molting hormone ecdysone, an ecdysteroid secreted by a pair of molting glands located in the body. Thus it appears that molting is triggered by a reduction in MIH in the blood, which stimulates the molting glands to synthesize and secrete ecdysone. It is thought that binding of MIH to a membrane receptor results in a cyclic nucleotide-dependent inhibition of ecdysone-synthesizing enzymes. Five genes were cloned in the tropical land crab that may play a role in MIH signaling: nitric oxide synthase (NOS), three guanylyl cyclases (GCs), and MIH. Proteomic approaches are used to identify and quantify levels and phosphorylation states of NOS and downstream targets of cyclic nucleotide-dependent protein kinases.

Regulation of muscle atrophy by myostatin. In mammals myostatin (Mstn) is a negative regulator skeletal muscle growth. We have recently identified Mstn orthologs in crabs and lobsters. As these genes are preferentially expressed in skeletal muscle, they may have similar functions in invertebrates. We have begun to examine the role of Mstn in controlling molt-induced claw muscle atrophy, in which the claw muscle mass is reduced as much as 78% before molting. The reduced mass facilitates withdrawal of the large claws through the narrow joints that connect the appendage to the body. Binding of Mstn to a membrane receptor results in phosphorylation of Smad proteins. Smads are transcription factors that translocate to the nucleus and alter gene expression. cDNAs encoding land crab Smads have been obtained. Quantitative RT-PCR and proteomic analysis will be used to measure Smad mRNA and protein levels, respectively. Image analysis will be used to quantify Smads in the nucleus and cytoplasm in response to Mstn.

References:

Kim, H.-W., S.G. Lee and D.L. Mykles. 2005. Ecdysteroid-responsive genes, RXR and E75, in the tropical land crab, Gecarcinus lateralis: differential tissue expression of multiple RXR isoforms generated at three alternative splicing sites in the hinge and ligand-binding domains. Molec. Cell. Endocrinol. 242:80-95.

Lee, S.G. and D.L. Mykles. 2006. Proteomics and signal transduction in the crustacean molting gland. Integr. Comp. Biol. 46:965-977.

Medler, S., T.R. Lilley, J.H. Riehl, E.P. Mulder, E.S. Chang and D.L. Mykles. 2007. Myofibrillar gene expression in differentiating lobster claw muscles. J. Exp. Zool. 307A:281-295.

Lee, K.J., H.-W Kim, A.M. Gomez, E.S. Chang, J.A. Covi and D.L. Mykles. 2007. Molt-inhibiting hormone from the tropical land crab, Gecarcinus lateralis: cloning, tissue expression, and expression of biologically active recombinant peptide in yeast. Gen. Comp. Endocrinol. 150:505-513.

Lee, K.J., R.M. Doran and D.L. Mykles. 2007. Crustacean hyperglycemic hormone from the tropical land crab, Gecarcinus lateralis: cloning, isoforms, and tissue expression. Gen. Comp. Endocrinol. 154:174-183.

Lee, S. G., B. D. Bader, E.S. Chang and D.L. Mykles. 2007. Effects of elevated ecdysteriod on tissue expression of three guanylyl cyclases in the tropical land crab, Gecarcinus lateralis: possible roles of neuropeptide signaling in the molting gland. J. Exp. Biol. 210:3245-3254.

Lee, S.G., H.-W. Kim and D.L. Mykles. 2007. Guanylyl cyclases in the tropical land crab: Gecarcinus lateralis: cloning of soluble (NO-sensitive and -insensitive) and membrane receptor forms. Comp. Biochem. Physiol. 2D:332-344.

Medler, S., E.S. Change and D.L. Mykles. 2007. Muscle-specific calpain is localized in regions near motor endplates in differentiating lobster claw muscles. Comp. Biochem. Physiol. 148A:591-598.


Illustration: Close-up of the land crab (Gecarcinus lateralis)