Identification of EXPA4 as a key gene in cotton salt stress adaptation through transcriptomic and coexpression network analysis of root tip protoplasts

Background: Salinity stress impairs cotton growth and fiber quality. Protoplasts enable elucidation of early salt-responsive signaling. Elucidating crop tolerance mechanisms that ameliorate these diverse salinity-induced stresses is key for improving agricultural productivity under saline conditions.

Results: Herein, we performed transcriptome profiling of Gossypium arboreum root tips and root tips-derived protoplasts to uncover salt tolerance genes and mechanisms. Differentially expressed genes (DEGs) were significantly enriched in the plant hormone signal transduction and MAPK signaling pathways. Transcriptome based weighted gene coexpression network analysis (WGCNA) clustered 885 commonly differentially expressed genes into four distinct modules. Black and yellow modules were highly upregulated under salt treatment, containing hub genes integral to signaling and transport, highlighting their importance. Differential expression analysis revealed more dynamic changes in protoplasts, identifying key genes including the Ga-α-expansin 4 (GaEXPA4). Silencing of the GaEXPA4 gene through virus-induced gene silencing heightened cotton's sensitivity to salt stress, leading to increased wilting, elevated lipid peroxidation, and impaired antioxidant activity under salt conditions compared to controls.

Conclusion: These findings underscore the functional significance of GaEXPA4 in the salt stress response. Future research should focus on elucidating the precise mechanisms of putative salt tolerance genes like GaEXPA4 and evaluating the potential of signaling pathways, such as MAPK, for engineering enhanced salt resilience in cotton. Integrating multi-omics approaches could further expand the genetic resources available for improving cotton cultivation in saline environments.