GhECA8 positively regulates salt tolerance via maintaining intracellular calcium homeostasis in cotton

Calcium (Ca 2 ⁺) signaling plays a central role in plant responses to salt stress, and endoplasmic reticulum-type Ca 2 ⁺-ATPases (ECAs), members of the P-type IIA Ca 2 ⁺-ATPase family, contribute to cellular Ca 2 ⁺ homeostasis by mediating intracellular Ca 2 ⁺ transport. However, the functions of ECA genes in cotton salt tolerance remain poorly understood. In this study, 24 ECA family members were identified across four cotton species, and GhECA8 was selected for functional characterization because of its strong and sustained induction by salt stress. Virus-induced gene silencing (VIGS) of GhECA8 significantly increased salt sensitivity, as evidenced by severe leaf wilting and chlorosis and reduced biomass. GhECA8 -silenced plants exhibited altered Ca 2 ⁺ regulation, characterized by enhanced local net Ca 2 ⁺ influx but reduced salt-induced accumulation of total leaf Ca, together with an increased Na⁺/K⁺ ratio, excessive reactive oxygen species (ROS) accumulation, weakened antioxidant capacity and osmotic adjustment, and increased cellular and tissue damage. In addition, GhECA8 silencing enhanced the expression of endoplasmic reticulum stress-related genes and attenuated salt-induced abscisic acid (ABA) accumulation and ABA-associated transcriptional responses. Collectively, these results indicate that GhECA8 contributes to cotton salt tolerance and is associated with coordinated changes in Ca 2 ⁺ homeostasis, ion balance, redox regulation, osmotic adjustment, and ABA-associated stress responses, making it a candidate for further functional evaluation in cotton.