Genome-wide prediction of secretory proteins in Ceratocystis paradoxa and expression analysis of effectors during early infection

BACKGROUND: Ceratocystis paradoxa, a filamentous ascomycete and soil-borne wound pathogen, attacks various host plants such as Ananas comosus, Cocos nucifera and Saccharum officinarum. Secretory proteins are critical for phytopathogenic infection, yet related studies on C. paradoxa remain scarce. METHODS: Using WoLF PSORT, TMHMM, and big-PI Predictor, we predicted the secretory proteins in C. paradoxa. Carbohydrate-active enzymes and pathogenicity-related proteins were predicted by HMMER and PHI-base. Effectors were predicted by EffectorP v3.0. RT-qPCR was used to screen effectors highly expressed during early infection. RESULTS: In this study, 253 secretory proteins were predicted from its 6,931 annotated genomic proteins. These proteins vary from 61 to 1894 residues in length, with signal peptides ranging from 13 to 35 residues in length. 58.89% of the signal peptide cleavage sites follow the canonical "Ala-X-Ala" motif, a substrate recognized by signal peptidase I. Using HMMER software, 78 carbohydrate-active enzymes (CAZymes) were predicted, including 45 glycoside hydrolases, 24 auxiliary activities, 4 polysaccharide lyases, 3 carbohydrate esterases, 1 glycosyl transferase and 1 carbohydrate binding module. Additionally, 144 secretory proteins matched known entries in the PHI-base, suggesting potential roles in pathogenicity. We further predicted 56 candidate effectors, classified into 9 cytoplasmic and 47 apoplastic types. Notably, six effector proteins were highly expressed during early infection, and three of them were predicted to be CAZymes. CONCLUSIONS: This study systematically predicted 253 potential secretory proteins from C. paradoxa via multi-tool prediction, which may provide a comprehensive inventory of its pathogenicity-related secretome. These proteins potentially include 78 carbohydrate-active enzymes, 144 pathogenicity-related proteins, and 56 candidate effectors. Notably, six effectors highly expressed during early infection-three of which may have carbohydrate-active enzyme activity-could be key candidates involved in initial infection. Our findings may fill the gap in C. paradoxa secretory protein research, lay a potential foundation for elucidating its pathogenic mechanisms, and provide potential targets for controlling this soil-borne pathogen.