Updated project metadata. Abstract Lysine crotonylation (Kcr) is a recently identified post-translational modification (PTM) in the prokaryotic and eukaryotic organisms. However, the function of lysine-crotonylated proteins in response to abiotic stress in plants remains largely unknown. Here, we report the first global profiling of the Kcr proteome in common wheat, identifying 4,696 Kcr sites on 1,726 substrate proteins, participating in a wide variety of biological and metabolic pathways. Combination of transcriptome and proteome in a RIL (recombinant inbred line) population, Kcr proteome, and genome-wide association study, a candidate gene phosphoglycerate kinase (TaPGK) was identified to potentially affect on cold tolerance in common wheat. Using EMS-mutants, overexpressed transgenic wheat plants and CRISPR/Cas9-mediated know-out mutants, we demonstrated that TaPGK played a positively key role in regulation of plant cold tolerance. Moreover, TaPGK protein strongly interacted with sirtuin-like (TaSRT1). EMS-mutant tetraploid and overexpressed transgenic wheat experiments confirmed that TaSRT1 gene negatively regulated wheat cold tolerance. Immunoprecipitation demonstrated that TaSRT1 promoted the degradation of TaPGK by erasing Kcr. Additionally, RNA-sequencing results showed that overexpressed TaPGK significantly improved expression of peroxidase genes. It concluded that a large number of Kcr sites were present in wheat important proteins and TaPGK crotonylated by TaSRT1 played a key role in wheat response to cold stress through scavenging the accumulation of ROS.