A single-cell atlas of Exopalaemon carinicauda gills provides insights into pillar cells mediated response of low-salinity stress in crustaceans

The ion homeostasis of crustaceans farmed in coastal areas is significantly influenced by low-salinity stress. Gill serves as the primary osmoregulatory organ in crustaceans, mediating ion exchange and acid-base balance. However, the cellular mechanisms underlying gill responses to low-salinity stress remain poorly understood. This study integrated cell structural analysis with single-cell RNA sequencing (scRNA-seq) to characterize low-salinity stress responses in Exopalaemon carinicauda gills. Low-salinity exposure induced significant structural alterations in pillar cells. scRNA-seq identified 11 transcriptionally distinct cell populations, including two ionocyte subtypes (pillar cells and septal cells). Comparative analysis revealed a significant increase in the abundance of pillar cell subcluster3 following low-salinity stress, with its associated differentially expressed genes (DEGs) enriched in ion transport pathways, notably hydrogen ion transport. Weighted gene co-expression network analysis (WGCNA) further indicated upregulation of hub genes involved in pH regulation within pillar cells. Pseudotime trajectory analysis suggested that gill adaptation to low-salinity stress involves a specific differentiation trajectory (cell fate 1) originating from pillar cells. Key DEGs in pillar cells responding to low-salinity stress, including V(H⁺)-ATPases and cytoplasmic carbonic anhydrase, were primarily linked to acid-base regulation. Collectively, these results elucidate gill cell heterogeneity and reveal pillar cell-mediated regulatory mechanisms underlying crustacean adaptation to low-salinity stress.