Systematic characterization of the cyclin gene family in the Chinese mitten crab Eriocheir sinensis and functional analysis of representative cyclins in bacteria-induced hemocyte proliferation.

Cyclins are core regulators of cell-cycle progression and also participate in transcriptional regulation and cellular stress responses. However, the cyclin family has not been systematically characterized in the Chinese mitten crab Eriocheir sinensis, and its potential role in hemocyte proliferation during antibacterial responses remains unclear. In this study, 18 cyclin family members were identified from the E. sinensis genome and designated as EsCyclins (abbreviated as EsCyc plus the subtype letter). Comparative analyses showed that these family members are distributed across 13 chromosomes and can be assigned to the major cyclin subfamilies on the basis of phylogenetic relationships, conserved motifs, gene structures, and domain organization. Representative cyclins, including EsCycA, EsCycB, EsCycB3, EsCycD, and EsCycE, displayed conserved sequence features and similar predicted core three-dimensional structures relative to invertebrate and vertebrate homologs. Tissue expression analysis further showed that these genes are broadly expressed but tissue biased, with relatively high transcript levels in hepatopancreas, testis, and, depending on the gene, stomach or muscle. After stimulation with heat-killed Vibrio parahaemolyticus, representative EsCyclin genes showed dynamic, gene-specific temporal expression patterns in synchronized primary hemocytes. Functional assays demonstrated that siRNA-mediated knockdown of EsCycA, EsCycD, or EsCycE significantly reduced hemocyte proliferation under bacterial stimulation, as assessed by flow cytometry and EdU incorporation. Together, this study provides the first systematic characterization of the cyclin family in E. sinensis and identifies EsCycA, EsCycD, and EsCycE as positive regulators of bacteria-induced hemocyte proliferation in vitro. These findings extend our understanding of cyclin-family evolution in crustaceans and provide a foundation for investigating how cell-cycle regulators participate in crustacean antibacterial cellular responses.