Molt regulation in green and red color morphs of the crab, Carcinus maenas: gene expression of molt-inhibiting hormone signaling components

Abstract

Summary In decapod crustaceans, regulation of molting is controlled by the X-organ/sinus gland complex in the eyestalks. The complex secretes molt-inhibiting hormone (MIH), which suppresses production of ecdysteroids by the Y-organ (YO). MIH signaling involves NO and cGMP in the YO, which expresses NO synthase (NOS) and NO-sensitive guanylyl cyclase (GC-I). Molting can generally be induced by eyestalk ablation (ESA), which removes the primary source of MIH, or by multiple leg autotomy (MLA). In our work on Carcinus maenas, however, ESA has limited effects on hemolymph ecdysteroid titers and animals remain in intermolt by 7 days post-ESA, suggesting that adults are refractory to molt induction techniques. Consequently, the effects of ESA and MLA on molting and YO gene expression in C. maenas green and red color morphotypes were determined at intermediate (16 and 24 days) and long-term (~90 days) intervals. In intermediate-interval experiments, ESA of intermolt animals caused transient 2- to 4-fold increases in hemolymph ecdysteroid titers during the first 2 weeks. In intermolt animals, long-term ESA increased hemolymph ecdysteroid titers 4 to 5-fold by 28 days post treatment, but there was no late premolt peak (>400 pg/μl) characteristic of late premolt animals and animals did not molt by 90 days post-ESA. There was no effect of ESA and MLA on the expression of Cm-elongation factor 2 (EF2), Cm-NOS, the beta subunit of GC-I (Cm-GC-Iβ), a membrane receptor GC (Cm-GC-II), and a soluble NO-insensitive GC (Cm-GC-III) in green morphs. Red morphs were affected by prolonged ESA and MLA treatments, as indicated by large decreases in Cm-EF2, Cm-GC-II, and Cm-GC-III mRNA levels. ESA accelerated the transition of green morphs to the red phenotype in intermolt animals, indicating that molting and integument color changes are not necessarily coupled. ESA delayed molting in premolt green morphs, whereas intact and MLA animals molted by 30 days post treatment. There were significant effects on YO gene expression in intact animals; Cm-GC-Iβ mRNA increased during premolt and Cm-GC-III mRNA decreased during premolt and increased during postmolt. Cm-MIH transcripts were detected in eyestalk ganglia, brain, and thoracic ganglion from green intermolt animals and ESA had no significant effect on Cm-MIH mRNA levels in brain and thoracic ganglion. The data suggest that MIH in the brain and thoracic ganglion prevents molt induction in green ESA animals.