Chemosensory proteins in the CSP4 clade evolved as plant immunity suppressors before two suborders of plant-feeding hemipteran insects diverged

Chemosensory proteins (CSPs) are small globular proteins with hydrophobic binding pockets that have a role in detection of chemicals, regulation of development and growth and host seeking behaviour and feeding of arthropods. Here, we show that a CSP has evolved to modulate plant immune responses. Firstly, we found that the green peach aphid Myzus persicae CSP Mp10, which is delivered into the cytoplasm of plant cells, suppresses the reactive oxygen species (ROS) bursts to both aphid and bacterial elicitors in Arabidopsis thaliana and Nicotiana benthamiana. In contrast, other CSPs, including MpOS-D1, do not have this ROS suppression activity. Aphid RNA interference studies demonstrated that Mp10 modulates the first layer of the plant defence response, specifically the BAK1 pathway. Alignment of CSPs from multiple aphid species showed that Mp10 homologues uniquely have tyrosine (Y40) and tryptophan (W120) flanking the central binding region. Exchange of aromatic residues between Mp10 and MpOS-D1 showed a gain of ROS activity of MpOS-D1 and loss of this activity of Mp10. We identified Mp10 homologs in diverse plant-sucking insect species, including aphids, whiteflies, psyllids and leafhoppers, but not in other insect species, including blood-feeding hemipteran insects. Moreover, the positions of Y and W residues are conserved among these Mp10 homologs, which we found also suppress plant ROS. Together, these data and phylogenetic analyses provides evidence that an ancestral Mp10-like sequence acquired plant ROS suppression activity via gain-of-function mutations before the divergence of plant-sucking insect species over 250 million years ago.