Cross-Talk of Protein Expression and Lysine Acetylation in Response to TMV Infection in Nicotiana benthamiana.

Lysine acetylation (Kac), a reversible PTM, plays an essential role in various biological processes, including those involving metabolic pathways, pathogen resistance, and transcription, in both prokaryotes and eukaryotes. TMV, the major factor that causes the poor quality of Solanaceae crops worldwide, directly alters many metabolic processes in tobacco. However, the extent and function of Kac during TMV infection have not been determined. The validation test to detect Kac level and viral expression after TMV infection and Nicotinamide (NAM) treatment clarified that acetylation was involved in TMV infection. Furthermore, we comprehensively analyzed the changes in the proteome and acetylome of TMV-infected tobacco (Nicotiana benthamiana) seedlings via LC-MS/MS in conjunction with highly sensitive immune-affinity purification. In total, 2082 lysine-acetylated sites on 1319 proteins differentially expressed in response to TMV infection were identified. Extensive bioinformatic studies disclosed changes in acetylation of proteins engaged in cellular metabolism and biological processes. The vital influence of Kac in fatty acid degradation and alpha-linolenic acid metabolism was also revealed in TMV-infected seedlings. This study first revealed Kac information in N. benthamiana under TMV infection and expanded upon the existing landscape of acetylation in pathogen infection.

Comprehensive Proteomic Analysis of Lysine Ubiquitination in Seedling Leaves of Nicotiana tabacum.

Lysine ubiquitination, a widely studied posttranslational modification, plays vital roles in various biological processes in eukaryotic cells. Although several studies have examined the plant ubiquitylome, no such research has been performed in tobacco, a model plant for molecular biology. Here, we comprehensively analyzed lysine ubiquitination in tobacco (Nicotiana tabacum) using LC-MS/MS along with highly sensitive immune-affinity purification. In total, 964 lysine-ubiquitinated (Kub) sites were identified in 572 proteins. Extensive bioinformatics studies revealed the distribution of these proteins in various cellular locations, including the cytoplasm, chloroplast, nucleus, and plasma membrane. Notably, 25% of the Kub proteins were located in the chloroplast of which 21 were enzymatically involved in important pathways, that is, photosynthesis and carbon fixation. Western blot analysis indicated that TMV infection can cause changes in ubiquitination levels. This is the first comprehensive proteomic analysis of lysine ubiquitination in tobacco, illustrating the vital role of ubiquitination in various physiological and biochemical processes and representing a valuable addition to the existing landscape of lysine ubiquitination.