As a novel post-translational modification (PTM), lysine 2-hydroxyisobutyrylation (Khib) is considered to regulate gene transcriptional activities in eukaryotic cells; however, the functions of Khib-modified proteins in plants remain unknown. Here, we report that Khib is an evolutionarily-conserved PTM in wheat and its progenitors. A total of 3348 Khib sites on 1074 proteins are identified in common wheat (Triticum aestivum L.) by using affinity purification and mass spectroscopy of 2-hydroxyisobutyrylome. Bioinformatic data indicate that Khib-modified proteins participate in a wide variety of biological and metabolic pathways. Immunoprecipitation confirms that Khib-modified proteins are present endogenously. A comparison of Khib and other main PTMs shows that Khib-modified proteins are simultaneously modified by multiple PTMs. Using mutagenesis experiments and co-immunoprecipitation assays, we demonstrate that Khib on K206 of phosphoglycerate kinase (PGK) is a key regulatory modification for its enzymatic activity, and mutation on K206 affects the interactions of PGK with its substrates. Furthermore, Khib modification of low-molecular-weight proteins is a response to the deacetylase inhibitors nicotinamide and trichostatin. This study provides evidence to promote our current understanding of Khib in wheat plants, including the cooperation between Khib and its metabolic regulation.
Keywords: Co-immunoprecipitation; Common wheat; Lysine 2-hydroxyisobutyrylation; Post-translational modification; Proteomics.
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