IntroductionMitogen-activated protein kinases (MAPKs) are central to stress and innate immune signaling in fish, but their genomic composition and infection related transcriptional responses remain unclear in bighead carp (Hypophthalmichthys nobilis).MethodsWe performed a comparative genomic survey across 24 species and characterized MAPK family members in bighead carp based on conserved motifs, gene structure, and synteny. Tissue expression profiling, transcriptome sequencing, qRT-PCR, weighted gene co-expression network analysis (WGCNA), and single-cell RNA sequencing were used to investigate MAPK expression and immune responses following Aeromonas hydrophila challenge. Cell-cell communication and pseudotime analyses were further applied to dissect infection responsive myeloid populations.ResultsWe identified 382 mapk sequences across 24 species, including 15 high confidence mapk genes in bighead carp, which were classified into the ERK, JNK, and p38 subfamilies. These genes were constitutively expressed across six tissues, with mapk12b and mapk14b showing relatively high expression in spleen and head kidney. Following A. hydrophila challenge, mapk6 and mapk11 were significantly upregulated, as confirmed by transcriptomic and qRT-PCR analyses. WGCNA further showed that multiple infection related modules were enriched in mapk family members.
IntroductionMitogen-activated protein kinases (MAPKs) are central to stress and innate immune signaling in fish, but their genomic composition and infection related transcriptional responses remain unclear in bighead carp (Hypophthalmichthys nobilis).MethodsWe performed a comparative genomic survey across 24 species and characterized MAPK family members in bighead carp based on conserved motifs, gene structure, and synteny. Tissue expression profiling, transcriptome sequencing, qRT-PCR, weighted gene co-expression network analysis (WGCNA), and single-cell RNA sequencing were used to investigate MAPK expression and immune responses following Aeromonas hydrophila challenge. Cell-cell communication and pseudotime analyses were further applied to dissect infection responsive myeloid populations.ResultsWe identified 382 mapk sequences across 24 species, including 15 high confidence mapk genes in bighead carp, which were classified into the ERK, JNK, and p38 subfamilies. These genes were constitutively expressed across six tissues, with mapk12b and mapk14b showing relatively high expression in spleen and head kidney. Following A. hydrophila challenge, mapk6 and mapk11 were significantly upregulated, as confirmed by transcriptomic and qRT-PCR analyses. WGCNA further showed that multiple infection related modules were enriched in mapk family members. Single-cell RNA sequencing of 28,348 spleen cells identified 13 distinct cell types and revealed marked expansion and activation of Neutrophils-I cells and M1 macrophages, accompanied by increased mapk6/mapk11 expression and strong enrichment of interferon stimulated pathways. Cell-cell communication and pseudotime analyses further suggested that tnf-tnfrsf1b signaling and progressive upregulation of mapk6/mapk11 along myeloid lineages contribute to the early splenic antibacterial response.ConclusionThis study provides a genome to single-cell view of MAPK family organization and immune function in bighead carp, and identifies mapk6 and mapk11 as potential mediators of the early spleen response to bacterial infection.