ABSTRACT The co‐infection of maize chlorotic mottle virus (MCMV) and sugarcane mosaic virus (SCMV) causes maize lethal necrosis (MLN), which seriously affects the yield and quality of maize. Ubiquitination is one of the most important protein post‐translational modifications. However, the role of ubiquitination modification in regulating maize resistance to viral infection remains largely unknown. In this study, we found that the ubiquitination levels in SCMV‐ and/or MCMV‐infected maize plants were higher than that in the non‐infected maize plants. Ubiquitinome and proteome analyses of the above maize plants revealed that most down‐regulated differentially accumulated proteins that possessed up‐regulated lysine ubiquitination sites were mainly involved in photosynthesis, fructose and mannose metabolism, and glyoxylate and dicarboxylate metabolism. Functional analyses of three DAPs involved in glyoxylate metabolism demonstrated that silencing ZmGOX1 facilitated SCMV and MCMV single and co‐infection, while knockdown of ZmHPR1 or ZmHPR2 suppressed viral infections. Moreover, overexpression of ZmGOX1 and its mutants at Kub sites enhanced maize resistance to SCMV infection. We also found that exogenous application of sodium sulphide could up‐regulate the expression of ZmGOX1 and effectively inhibit viral infections. These findings provide novel insights into the roles of ubiquitination in the regulation of maize resistance to viral infection.