DTX3L-PARP9 co-evolve as a single adaptive unit linking ubiquitination and ADP-ribosylation in antiviral immunity

Functional protein complexes are central to innate immunity, by integrating and diversifying signaling pathways of immune responses. Understanding how such complexes evolve is therefore key to elucidate the molecular basis of immune adaptation. Using a comparative genomics framework, we show that the DTX3L-PARP complex, which integrates ubiquitin-dependent and ADP-ribosylation pathways to regulate antiviral immunity, is an ancient and rapidly evolving functional module in vertebrates. We demonstrate that this complex has undergone recurrent adaptive evolution, with strong signatures of co-evolution between its components, consistent with early functional coupling and coordinated evolution as a single adaptive unit rather than independent proteins. Adaptive residues are enriched at protein-protein interaction surfaces within the RING-DTC domain of DTX3L, while PARP9 and PARP14 concentrate adaptive changes in regions involved in complex assembly and catalytic activities, consistent with hallmarks of pathogen-driven selection. These evolutionary signals remain detectable at short evolutionary timescales, including within species and sub-species, indicating ongoing adaptive evolution of the complex. These patterns support a model in which pathogens recurrently target the DTX3L-PARP axis to disrupt ubiquitin-ADP-ribose signaling, either by simultaneously interfering with catalytic domains or by destabilizing the complex interfaces. Altogether, our findings reveal the DTX3L-PARP9-PARP14 complex as a co-evolving adaptive module shaped by persistent host-virus arms races, highlighting how interconnected post-translational modification systems evolve in concert to sustain vertebrate antiviral immunity.