Comprehensive analysis of colony-stimulating factor genes and immunostimulatory effects of CSF3a in rainbow trout (Oncorhynchus mykiss).

Colony-stimulating factors (CSFs) play critical roles in the host response to tissue injury and pathogenic infection. In this study, we conducted a comprehensive genome-wide analysis and identified seven csf genes in rainbow trout (Oncorhynchus mykiss). Comparative analyses of gene structure, conserved synteny, and protein sequence alignment further supported their annotation and orthology with other vertebrate csf genes. Although csf3 has been identified in several teleost species, its biological functions remain largely uncharacterized. Here, we reported that rainbow trout csf3 (Rtcsf3) genes exhibited broad tissue distribution and were significantly upregulated in response to Aeromonas salmonicida infection and pathogen analog stimulation, suggesting their potential role in innate immunity. The open reading frame (ORF) of csf3a encodes a 215-amino acid (aa) precursor protein containing a signal peptide and a conserved IL-6-type cytokine domain. Functional assays demonstrated that recombinant CSF3a (rRtCSF3a) significantly increased the expression of inflammatory cytokines (il1β, il10 and tnfα) and promoted splenic cell proliferation in vitro. Furthermore, transcriptomic profiling of rRtCSF3a-stimulated spleen cells revealed enrichment of genes associated with cytokine signaling, cell proliferation, programmed cell death, and metabolic reprogramming. Collectively, these findings provide the first comprehensive characterization of the csf gene family in teleosts and uncover the immunostimulatory and regulatory functions of CSF3a in rainbow trout.