A functional VCBP-C1q interaction in amphioxus reveals evolutionary origins of the classical complement pathway.

The origin of the classical complement pathway remains unclear, primarily due to the evolutionary interplay between immunoglobulin (Ig) and C1q. An Ig superfamily member, variable region-containing chitin-binding protein 5 (VCBP5), from the basal chordate amphioxus exhibited a marked upregulation upon E. coli stimulation and accumulated in the epithelial lining and circulatory sinuses of the hepatic caecum. Recombinant VCBP5 (rVCBP5) exhibits strong binding affinity for E. coli via the C'C'' loop of its IgV2 domain. Exogenous addition of rVCBP5 not only enhances the bactericidal activity of humoral fluids but also provides robust protection against bacterial proliferation in vivo. Notably, rVCBP5 specifically binds to the gC1q domain of BjC1q, with the interdomain hinge regions of VCBP5 serving as key interaction interfaces. Moreover, rBjC1q directly interacts with the serine protease BjMASP1/3. Importantly, formation of the VCBP5-bacterial complex facilitates rBjC1q-mediated recruitment of BjMASP1/3, thereby triggering complement activation. Our findings uncover a functional VCBP-C1q partnership in amphioxus, revealing a primitive complement activation pathway. This discovery provides new insights into the evolutionary emergence of the classical complement pathway.