The Rap subfamily in Mugil cephalus: genome-wide identification and a role for McRap1a in antibacterial immunity.

Abstract

The Rap subfamily of small GTPases is crucial signaling nodes in eukaryotes, but their immune functions in teleost fish remain incompletely characterized. In this study, we performed genome-wide identification of the Rap genes in flathead grey mullet (Mugil cephalus) and functionally characterized one member, McRap1a, using bioinformatics and recombinant protein assays. Five Rap members (McRap1a, McRap1b, McRap2a, McRap2b, and McRap2c) were identified, all showing high evolutionary conservation with vertebrate orthologs. Mcrap1a was expressed in all tested tissues, with the highest transcript abundance in the head kidney, and showed tissue- and time-dependent responses to lipopolysaccharide challenge, including significant upregulation in the gill and spleen. Recombinant McRap1a protein directly bound to both Gram-negative and Gram-positive bacteria, exerted concentration-dependent antibacterial activity with particular potency against pathogenic Vibrio species, inhibited biofilm formation across all tested strains, and caused selective bacterial membrane disruption. These findings establish McRap1a as a direct innate immune effector in mullet, providing the functional evidence that a teleost Rap protein possesses broad antimicrobial and anti-biofilm activities, and offering potential targets for aquaculture disease control.