Identification of a Snf7-domain-containing protein that exhibits high affinity and synergistic activity for Cry13Aa1 toxin in Bursaphelenchus xylophilus

Abstract

Pine wilt disease, caused by the pinewood nematode Bursaphelenchus xylophilus (Rhabditida: Aphelenchoididae), results in significant global economic and ecological impacts. Although the Cry13Aa1 toxin from Bacillus thuringiensis shows nematicidal activity, its mechanism of action against B. xylophilus remains unclear. This study aimed to identify and characterize the receptors for Cry13Aa1 in B. xylophilus. We cloned the cDNAs encoding an Snf7 domain-containing protein (BxSnf7) from B. xylophilus. Far-western blot analysis revealed a specific binding interaction between BxSnf7 and Cry13Aa1, showing a dissociation constant (K_d) of 20.8 ± 4.2 nM. Interestingly, bioassay results indicated that silencing BxSnf7 increased the susceptibility of nematodes to Cry13Aa1 at higher concentrations, although the difference was not statistically significant. Besides, the combined application of BxSnf7 with Cry13Aa1 significantly enhanced nematicidal mortality (95.9 %) after 24 h of treatment, which higher than the expected mortality (42.8 %) (χ² = 16.118, P = 0.048), indicating that the exogenous BxSnf7 synergistically enhances the activity of Cry13Aa1 toxin. These findings identify BxSnf7 as a novel Cry13Aa1 binding protein and reveal a unique mechanism by which BxSnf7 synergistically enhances the activity of Cry13Aa1. However, BxSnf7 does not function as the primary receptor, and further research is needed to investigate its role in modulating nematode susceptibility to Cry13Aa1.