<p>In crustaceans, 20-hydroxyecdysone (20E), the primary bioactive form of ecdysteroid hormones, regulates molting and ovarian development. <i>In vivo</i> and <i>in vitro</i> approaches were used to examine the potential nongenomic mechanisms through which 20E modulates ovarian maturation in Chinese mitten crabs (<i>Eriocheir sinensis</i>) and its effects on signaling components and associated genes within the phospholipase C (PLC)-protein kinase C (PKC) pathway. The results demonstrate that the administration of 20E considerably upregulated the hepatopancreatic mRNA expression of G proteins (<i>Gs</i> and <i>Gq</i>), calmodulin-dependent protein kinase II, and inositol 1,4,5-trisphosphate receptors. Concurrent increases in phosphatidylinositol-specific PLC activity were observed, along with elevated levels of its catalytic products (<i>i.e.</i>, inositol 1,4,5-trisphosphate and diacylglycerol). Moreover, 20E stimulation activated the phosphorylation and enhanced the transcriptional expression of PKC. Notably, <i>in vitro</i> pharmacological inhibition with U73122 (a PLC inhibitor) and Go6983 (a PKC inhibitor) resulted in the effective suppression of 20E-induced expression of downstream target genes, including the ecdysone receptor (<i>EcR</i>), retinoid X receptor (<i>RXR</i>), vitellogenin (<i>Vg</i>), and Vg receptor (<i>VgR</i>). These findings suggest that 20E modulates hepatopancreatic Vg synthesis in the Chinese mitten crab through PLC-PKC signaling transduction, thereby exerting indirect regulatory effects on ovarian development.</p>