Psyllid Cysteine Cathepsins Directly Cleave the Outer Membrane Protein BamD of Citrus Huanglongbing Pathogen

Candidatus Liberibacter asiaticus (CLas), the causative agent of citrus Huanglongbing (HLB), is transmitted by Asian citrus psyllid, Diaphorina citri; however, the role of the insect's immune effectors in defending against CLas remains poorly understood. This study reveals that D. citri cathepsin L (DcCTSL1), a member of the cysteine protease family, serves as a key immune effector directly cleaving CLas outer membrane protein BamD, thereby defending against CLas infection in insect vectors. Upon CLas infection, DcCTSL1 expression is upregulated. This protease targets the CLas membrane by specifically interacting with BamD, which in turn enables DcCTSL1 to recognise the conserved Ile145-Arg146 motif of BamD. Following this recognition, DcCTSL1 cleaves the peptide bond between Arg146 and Asp147 of BamD, an action that is mechanistically dependent on the conserved catalytic triad (Cys185, His325, and Asn350) of DcCTSL1. Heterologous application of DcCTSL1 in citrus plants confers resistance to CLas. Furthermore, a homologous citrus cysteine protease also cleaves BamD via the same mechanism. Our study reveals an evolutionarily conserved defence mechanism mediated by cysteine proteases against CLas in both insect vectors and plant hosts, thus offering a novel foundation for developing protease-based strategies against HLB.