The involvement of Rab5 in regulating haematopoiesis in the Chinese mitten crab Eriocheir sinensis.

Rab5 functions as a pivotal regulator in the intricate processes of membrane trafficking, orchestrating a multitude of cellular activities. In the present study, a Rab5 homolog with conserved structure features was identified from Chinese mitten crab Eriocheir sinensis (designated EsRab5). The mRNA transcripts of EsRab5 were detected in all the tested tissues, with particularly high expression levels observed in brain and haematopoietic tissue (HPT). Notably, its mRNA expression in HPT was significantly up-regulated at 6 and 12 h following stimulation with Aeromonas hydrophila. Immunocytochemical assay showed that EsRab5 protein was diffusely distributed throughout the HPT, with a particularly prominent concentration in the cytoplasm. After A. hydrophila stimulation, the immunoreactive signals for EsRab5 in HPT were markedly more intense compared to those in the control group. Upon injection of EsRab5-specific siRNA to inhibit its expression, a significant increase in the percentage of EdU-positive cells within HPT was observed following A. hydrophila stimulation, which was 2.62-fold (p < 0.0001) of that in the EGFP-RNAi group. Meanwhile, the expression levels of proliferation related factors (EsRunx, EsGLP and EsAstakine), cell cycle-related proteins (EsCyclin E, EsCDK2, EsCDK4, and EsCyclin D) as well as the MAPK signal pathway were increased significantly in EsRab5-RNAi crabs after A. hydrophila stimulation. These results suggested that EsRab5 serves as a critical regulator in homeostasis maintenance of haematopoiesis in E. sinensis by modulating multiple factors.