Comparative secretomics identifies conserved WAxR motif-containing effectors in rust fungi that suppress cell death in plants

Identification of novel effectors with conserved features has always remained a challenge in plant-pathogen interaction studies. The introduction of the genomics era in plant-pathogen studies has led to the identification of significant candidate effectors with novel motifs such as RxLR and dEER motifs. However, in the case of fungal pathogens, limited conserved motifs associated with effectors have been discovered yet. In the present study, we have performed comparative secretome analysis for major plant pathogens of diverse nutrition mechanisms with the aim of dissecting the features underlying their corresponding secretome and conserved motifs. We showed that rust fungi possess the lowest Cell wall degrading enzymes (CWDEs) consortium lower than other biotrophic pathogens. We also showed rust fungi possess the highest secretory superoxide dismutase (SOD) than other studied plant pathogens. Further, we prioritized the candidate secretory effectors proteins (CSEPs) of all the studied pathogens by combining various effector mining parameters to highlight the candidates with potential effector features. A novel WAxR motif in conjugation with the Y/F/WxC (FGC) motif was identified in the effectors of various P. striiformis races present globally. The WAxR/WAxR like motifs ( WxxR, WAxx, xAxR) containing effectors were also found in the secretome of other rust fungi. Further, the functional validation of two candidate effectors with WAxR motif from P. striiformis Yr9 showed that these effectors localize to the nucleus as well as cytoplasm, and are able to suppress BAX induced cell death in Nicotiana benthamiana. The mutation analysis of individual residues of the WAxR motif (W, A, R) however did not affect the cell death suppression nor subcellular localization of these effectors. Overall, the current study reports the presence of novel motifs in large numbers of effectors of rust fungi with cell death suppression features. HighlightsO_LISecretome analysis of various plant pathogens performed C_LIO_LIA prioritization list for candidate effectors designed C_LIO_LIA novel WAxR motif was discovered in rust fungal effector proteins C_LIO_LITwo WAxR candidate effectors were functionally validated C_LI