LEE, SG*; KIM, HW; BADER, BD; CHANG, ES; MYKLES, DL; Colorado State Univ, Fort Collins, CO; Univ. of Utah, Salt Lake City, UT; Colorado State Univ, Fort Collins, CO; Bodega Marine Lab, Bodega Bay, CA; Colorado State Univ, Fort Collins, CO: Effects of elevated ecdysteroid on expression of three guanylyl cyclases in tropical land crab, Gecarcinus lateralis

Guanylyl cyclases (GCs) catalyze the production of the second messenger, cyclic guanosine monophosphate (cGMP) from GTP. cGMP is a key signaling molecule that activates cGMP-dependent kinases, which leads to metabolic changes in target tissues. Increased cGMP is associated with inhibition of ecdysteroid secretion by the crustacean molting gland or Y-organ (YO). Moreover, NO synthase is expressed in the YO, suggesting that NO-sensitive GC is a component of the signaling pathway regulating ecdysteroid synthesis. Four GC classes occur in insects: NO-sensitive soluble GC (GC-I), membrane GC (GC-II), and two NO-insensitive soluble GCs (GC-III & GC-IV). cDNAs encoding the β subunit of GC-I, GC-II, and GC-III were cloned from land crab. Two GC-Iβ isoforms differing in the absence (+0N) or presence (+32N) of a 32-amino acid sequence at the N-terminus were characterized. Three GC-II isoforms, which varied in length within the ligand-binding domain (+18, +9, and +0 amino acid insertions), differed in tissue expression: GC-II⁺¹⁸ was expressed only in heart and skeletal muscle; GC-II⁺⁹ was expressed in all tissues; and GC-II⁺⁰ was expressed in most tissues. GC-II⁺⁹ is the putative receptor for crustacean hyperglycemic hormone (CHH). Increased ecdysteroid induced by eyestalk (ES) ablation increased GC-Iβ and GC-III mRNA levels in the YO, but had no effect on the mRNA levels of the GC-II⁺⁰ and GC-II⁺⁹ isoforms. These data suggest that tissues differ in their response to CHH and related neuropeptides, and that GC-Iβ and GC-III expression is stimulated by decreased ES neuropeptides and/or increased ecdysteroids in the hemolymph. Experiments in which 20-hydroxyecdysone is injected into intact animals will determine if GC-Iβ and GC-III mRNA levels are directly regulated by ecdysteroid in YO and other tissues. Supported by NSF (IBN-0342982).