Impact of alternative splicing on Arabidopsis proteome (Part III, TMT RT-SPS-MS3)

ABSTRACT: Alternative splicing (AS) is a key regulatory process in eukaryotes. Despite its importance, the impact of AS on plant protein diversity and its global effect on the proteome remain largely unexplored. To address this, we included the AS acinus pinin mutant in our qualitative proteomic analysis to identify splice variants, and in our quantitative analysis to assess the impact of intron retention (IR) on protein abundance. Specifically, we (1) investigated global proteomic changes in acinus pinin double mutants using tandem mass tag (TMT) labeling. ACINUS and PININ are evolutionarily conserved splicing factors, and their disruption induces splicing events resembling those triggered by various stressors. We employed TMT labeling with real-time search MS3 (RTS-SPS-MS3), combined with extensive sample fractionation, to maximize proteome coverage and achieve accurate quantification. (2) To enhance isoform-specific peptide detection, we incorporated data generated using the alternative protease AspN, along with mined data from the Arabidopsis Proteome Draft (Mergner et al., 2020, Nature). This TMT-based approach allowed us to assess how increased intron retention (IIR) affects the proteome by integrating proteomic data with matched transcriptomic profiles. Moreover, our method enabled identification of isoform-specific peptides—both annotated and previously unannotated—using in-house algorithms and filtering steps. These findings demonstrate that alternatively spliced transcripts are translated and detectable at the protein level, providing insight into the functional consequences of isoform diversity and revealing the complex relationship between AS and protein abundance.

INSTRUMENT(S):

ORGANISM(S): Arabidopsis Thaliana (mouse-ear Cress)

TISSUE(S): Whole Body

SUBMITTER: Shouling Xu  

LAB HEAD: Shouling Xu

PROVIDER: PXD068593 | Pride | 2026-06-08

REPOSITORIES: Pride