Four-Dimensional Data-Independent Acquisition-Based Proteomic Profiling Combined with Transcriptomic Analysis Reveals...

In green plants, the chloroplast is responsible for light energy transition and organic assimilation. However, the molecular mechanisms underlying chloroplast development in horticultural crops remain unclear. Here, four-dimensional data-independent acquisition-based proteomic profiling identified 1,727 differentially expressed proteins between "Zhongshu 4" (ZS4) and yellowing mutant (ym) leaves, a considerable proportion of which were down-regulated chloroplast proteins. Functional analysis revealed that light harvesting and chlorophyll biosynthesis were correlated with ym leaf yellowing, validated by RNA sequencing. Quantitative PCR confirmed that chlorophyll a/b-binding protein 3C (CAB3C) related to light harvesting and NADPH:protochlorophyllide oxidoreductase 3 (POR3) involved in chlorophyll biosynthesis were repressed in ym leaves. Virus-induced gene silencing showed that suppressing CAB3C and POR3 decreased the net photosynthetic rate and chlorophyll content. Additionally, the F-box protein Slym1 negatively regulated the expression of CAB3C by depressing transcription factor SlFHY3 levels. Our findings offer insights into the regulatory mechanisms of chloroplast development in tomato.