Disease-Resistance Functional Analysis and Screening of Interacting Proteins of ZmCpn60-3, a Chaperonin 60 Protein from Maize

Chaperonin 60 proteins plays an important role in plant growth and development as well as the response to abiotic stress. As part of the protein homeostasis system, molecular chaperones have attracted increasing attention in recent years due to their involvement in the folding and assembly of key proteins in photosynthesis. However, little is known about the function of maize chaperonin 60 protein. In the study, a gene encoding the chaperonin 60 proteins was cloned from the maize inbred line B73, and named <i>ZmCpn60-3</i>. The gene was 1, 818 bp in length and encoded a protein consisting of 605 amino acids. Phylogenetic analysis showed that ZmCpn60-3 had high similarity with OsCPN60-1, belonging to the β subunits of the chloroplast chaperonin 60 protein family, and it was predicted to be localized in chloroplasts. The <i>ZmCpn60-3</i> was highly expressed in the stems and tassels of maize, and could be induced by exogenous plant hormones, mycotoxins, and pathogens; Overexpression of <i>ZmCpn60-3</i> in <i>Arabidopsis</i> improved the resistance to <i>Pst</i> DC3000 by inducing the hypersensitive response and the expression of SA signaling-related genes, and the H₂O₂ and the SA contents of <i>ZmCpn60-3</i>-overexpressing <i>Arabidopsis</i> infected with <i>Pst</i> DC3000 accumulated significantly when compared to the wild-type controls. Experimental data demonstrate that flg22 treatment significantly upregulated transcriptional levels of the PR1 defense gene in <i>ZmCpn60-3</i>-transfected maize protoplasts. Notably, the enhanced resistance phenotype against Pseudomonas syringae pv. tomato DC3000 (<i>Pst</i> DC3000) in <i>ZmCpn60-3</i>-overexpressing transgenic lines was specifically abolished by pretreatment with ABT, a salicylic acid (SA) biosynthetic inhibitor. Our integrated findings reveal that this chaperonin protein orchestrates plant immune responses through a dual mechanism: triggering a reactive oxygen species (ROS) burst while simultaneously activating SA-mediated signaling cascades, thereby synergistically enhancing host disease resistance. Additionally, yeast two-hybrid assay preliminary data indicated that ZmCpn60-3 might bind to ZmbHLH118 and ZmBURP7, indicating ZmCpn60-3 might be involved in plant abiotic responses. The results provided a reference for comprehensively understanding the resistance mechanism of <i>ZmCpn60-3</i> in plant responses to abiotic or biotic stress.