Milograna Sarah Ribeiro, Bell Fernanda Tinti, McNamara John Campbell
Departamento de Biologia, FFCLRP, Universidade de São Paulo, Ribeirão Preto, 14040-901 São Paulo, Brazil.
Biol Bull. 2012 Oct;223(2):178-91. doi: 10.1086/BBLv223n2p178.
Crustacean color change results partly from granule aggregation induced by red pigment concentrating hormone (RPCH). In shrimp chromatophores, both the cyclic GMP (3', 5'-guanosine monophosphate) and Ca(2+) cascades mediate pigment aggregation. However, the signaling elements upstream and downstream from cGMP synthesis by GC-S (cytosolic guanylyl cyclase) remain obscure. We investigate post-RPCH binding events in perfused red ovarian chromatophores to disclose the steps modulating cGMP concentration, which regulates granule translocation. The inhibition of calcium/calmodulin complex (Ca(2+)/CaM) by N-(6-aminohexyl)-5-chloro-1-naphthalenesulphonamide (W7) induces spontaneous aggregation but inhibits RPCH-triggered aggregation, suggesting a role in pigment aggregation and dispersion. Nitric oxide synthase inhibition by Nω-nitro-L-arginine methyl ester hydrochloride (L-NAME) strongly diminishes RPCH-induced aggregation; protein kinase G inhibition (by rp-cGMPs-triethylamine) reduces RPCH-triggered aggregation and provokes spontaneous dispersion, disclosing NO/PKG participation in aggregation signaling. Myosin light chain phosphatase inhibition (by cantharidin) accelerates RPCH-triggered aggregation, whereas Rho-associated protein kinase inhibition (by Y-27632, H-11522) reduces RPCH-induced aggregation and accelerates dispersion. MLCP (myosin light chain kinase) and ROCK (Rho-associated protein kinase) may antagonistically regulate myosin light chain (MLC) dephosphorylation/phosphorylation during pigment dispersion/aggregation. We propose the following general hypothesis for the cGMP/Ca(2+) cascades that regulate pigment aggregation in crustacean chromatophores: RPCH binding increases Ca(2+)(int), activating the Ca(2+)/CaM complex, releasing NOS-produced nitric oxide, and causing GC-S to synthesize cGMP that activates PKG, which phosphorylates an MLC activation site. Myosin motor activity is initiated by phosphorylation of an MLC regulatory site by ROCK activity and terminated by MLCP-mediated dephosphorylation. Qualitative comparison reveals that this signaling pathway is conserved in vertebrate and invertebrate chromatophores alike.
甲壳类动物的颜色变化部分是由红色素浓缩激素（RPCH）诱导的颗粒聚集引起的。在虾的色素细胞中，环鸟苷酸（3',5'-鸟苷单磷酸）和Ca(2+)信号通路都介导色素聚集。然而，GC-S（胞质鸟苷酸环化酶）合成cGMP上游和下游的信号元件仍不清楚。我们研究了灌注的红色卵巢色素细胞中RPCH结合后的事件，以揭示调节cGMP浓度的步骤，而cGMP浓度调节颗粒转运。N-(6-氨基己基)-5-氯-1-萘磺酰胺（W7）对钙/钙调蛋白复合物（Ca(2+)/CaM）的抑制诱导了自发聚集，但抑制了RPCH触发的聚集，表明其在色素聚集和分散中起作用。盐酸Nω-硝基-L-精氨酸甲酯（L-NAME）对一氧化氮合酶的抑制强烈减弱了RPCH诱导的聚集；蛋白激酶G抑制（通过rp-cGMPs-三乙胺）降低了RPCH触发的聚集并引发自发分散，揭示了NO/PKG参与聚集信号传导。肌球蛋白轻链磷酸酶抑制（通过斑蝥素）加速了RPCH触发的聚集，而Rho相关蛋白激酶抑制（通过Y-27632、H-11522）降低了RPCH诱导 的聚集并加速了分散。MLCP（肌球蛋白轻链激酶）和ROCK（Rho相关蛋白激酶）可能在色素分散/聚集过程中对肌球蛋白轻链（MLC）的去磷酸化/磷酸化起拮抗调节作用。我们提出了以下关于调节甲壳类动物色素细胞中色素聚集的cGMP/Ca(2+)信号通路的一般假设：RPCH结合增加Ca(2+)(int)，激活Ca(2+)/CaM复合物，释放一氧化氮合酶产生的一氧化氮，并使GC-S合成激活PKG的cGMP，PKG使MLC激活位点磷酸化。肌球蛋白运动活性由ROCK活性使MLC调节位点磷酸化启动，并由MLCP介导的去磷酸化终止。定性比较表明，该信号通路在脊椎动物和无脊椎动物的色素细胞中都是保守的。
