Wheat (Triticum aestivum L.) is an important staple crop for global human.  The necrotrophic fungus Rhizoctonia cerealis is the causal pathogen of sharp eyespot, a devastating disease of wheat.  Herein, we identified RcMEP1, a zinc metalloprotease-encoding gene from R. cerealis genomic sequences, and characterized its pathogenesis function.  RcMEP1 expressed at markedly-high levels during R. cerealis infection process to wheat.  The predicted protein RcMEP1 comprises of 287 amino acid residues and contains a signal peptide and a M43 metalloprotease domain harboring the active site motif (HEVGHWLGLYH).  The assays of Agrobacterium tumefaciens-mediated transient expression in Nicotiana benthamiana leaves indicated that RcMEP1 is an apoplastic elicitor of cell death, and that the predicted signal peptide functions and is required for secretion and cell death-induction.  The purified RcMEP1 protein and its M43 domain peptide were individually able to induce plant cell death and H₂O₂ accumulation, and to inhibit expression of host chitinases when infiltrated into wheat and N. benthamiana leaves, while the M43 domain-deleting peptide and negative control lacked the capacity.  Moreover, compared with the control pretreatment, the purified RcMEP1 protein or its M43-domain peptide resulted in enhanced pathogenesis in the inoculated wheat, whereas the M43 domain-deleting peptide failed.  These results suggest that RcMEP1 acted as an important pathogenicity factor during R. cerealis infection to wheat and that its signal peptide and M43 domain are required for the secretion and pathogenesis of RcMEP1.  This study provides insights into pathogenesis role of M43 domain-containing metalloproteases during R. cerealis infection to wheat.

 

Receptor-like cytoplasmic kinases (RLCKs) represent a large family of proteins in plants.  In Arabidopsis and rice, several RLCKs in subfamily VII (RLCKs-VII) have been implicated in pathogen-associated molecular pattern-triggered immunity and basal resistance against bacterial and fungal pathogens.  However, little is known about roles of RLCKs-VII of the important crop common wheat (Triticum aestivum) in immune responses.  Here, we isolated a RLCK-VII-encoding gene from wheat, designated as TaRLCK1B, and investigated its role in host immune response to infection of a necrotrophic fungus Rhizoctonia cerealis that is a major pathogen of sharp eyespot, a destructive disease of wheat.  RNA-sequencing and RT-qPCR analyses showed that transcriptional level of TaRLCK1B was significantly higher in sharp eyespot-resistant wheat cultivars than in susceptible wheat cultivars.  The gene transcription was rapidly and markedly elevated in the resistant wheat cultivars by R. cerealis infection.  The TaRLCK1B protein was closely related to OsRLCK176, a rice resistance-related RLCKs-VII, with 84.03% identity.  Virus-induced gene silencing plus wheat response to R. cerealis assay results indicated that silencing of TaRLCK1 impaired resistance to R. cerealis.  Meantime, silencing of TaRLCK1 significantly elevated both the content of H₂O₂ (a major kind of reactive oxygen species, ROS) and the transcriptional level of the ROS-generating enzyme-encoding gene RBOH, but repressed the expression of the ROS-scavenging enzyme-encoding gene CAT1 at 18 hours after inoculation (hai) with R. cerealis.  Taken together, these data suggested that TaRLCK1B was required for the early immune response of wheat to R. cerealis through modulating ROS signaling in wheat.