Dataset Information

Impact of alternative splicing on Arabidopsis proteome (Part I, AspN)

ABSTRACT: Alternative splicing (AS) is a key regulatory process in eukaryotes. Despite its importance, the impact of AS on proteome diversity in plants remains largely unexplored. Traditional tryptic digestion has limitations in detecting junction peptides because evolutionarily conserved nucleotide usage at exon boundaries increases the frequency of lysine- and arginine-coding triplets at exon ends, making detection of exon–exon and exon–intron junctions challenging. To address this gap, we performed a comprehensive analysis of the Arabidopsis proteome using the protease AspN combined with extensive offline fractionation. AspN is an endoprotease that cleaves at the N-terminal side of aspartic acid residues. To further enhance detection of alternatively spliced proteins, we analyzed both wild-type plants and the AS mutant acinus pinin. This approach enabled identification of isoform-specific peptides—both annotated and previously unannotated—using in-house algorithms and rigorous filtering steps. Our findings demonstrate that alternative splicing contributes substantially to proteome diversity in plants.

INSTRUMENT(S):

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

TISSUE(S): Whole Body

SUBMITTER: Shouling Xu

LAB HEAD: Shouling Xu

PROVIDER: PXD068586 | Pride | 2026-06-08

REPOSITORIES: Pride

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Impact of alternative splicing on Arabidopsis proteome.

Reyes Andres V AV Zhang Christopher C Karunadasa Sumudu S SS Shrestha Ruben R Grismer TaraBryn S TS Byun Danbi D Xu Shou-Ling SL

Nucleic acids research 20260401 8

Limited proteomic evidence makes it unclear to what extent alternative splicing (AS) isoforms are translated and functionally relevant in eukaryotes. Here, we present a comprehensive proteomic analysis in plants using large-scale data mining, extensive fractionation of AspN- and trypsin-digested proteomes, and both label-free and TMT labeling. In total, we identified 471 196 peptides from 22 479 proteins by searching against Araport11, revealing 32 110 isoform-specific peptides. Using an integra ...[more]

PMID: 42080264

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Project description:Light is a pivotal signal for plants’ growth and survival. The promptly responses are required and achieving by sophisticated transcriptomic adjustments, the promptly translation enhancement, and effective protein surveillance. However, the global role of alternative splicing regulations in response to the light is unknown. Through global surveys of alternative splicing profile and frequency calculation for individual events normalized to transcriptomic changes, we identify nearly half of transcript present at this stage are alternatively spliced. Through the unbiased event comparison in D and L4h mRNA, we reveal a clear impact of splicing control on 2,961 genes during early photomorphogenesis. Intron retention is the most frequently event under controls of light as discovered in other developmental tissues and stages of Arabidopsis. Cryptic exon is the most abundant event preserved only in the dark-grown Arabidopsis. Genes underlying the splicing and transcriptional control mostly not overlapped indicate the distinct impacts of two regulatory mechanisms. Through confirmation with a homemade microarray for their responsiveness of light, we conclude 1,792 genes as biological validated candidates. Through functional study for genes underlying light-mediated splicing controls, we explore new light-signaling components which mainly responsive in isoform abundance but steady state mRNA. Among examination both positive (RPR39, KH-RBP, RRM and RRC1) and negative (PIF4, AMI1, DNAJ, UKL3, TF-IIs-like, and ubknown-Golgi protein) factors are controlled by splicing. Through predictions for example isoform structures, PIF4-IR4, AMI1-IR7-IR8, and DNAJ-IR1 both possess PTC and possibly produce truncated coding protein. PIF4-IR4 becomes best illustration for the dysfunction for the negative regulator by light. Above PTC+ isoforms together with one third of retained introns translate that evident for NMD-insensitive retained introns and supplements for protein levels. The event identities and cis-motif signatures associated with light regulations confirm a given uniqueness for the gene expression regulation during Arabidopsis photomorphogenesis. mRNA and polysome-associated mRNA of 4 days old Arabidopsis seedling grown under the dark or with 4 h light-treatment were isolated and the transcriptome and splicing events profiles were surveyed by mRNAseq. Independent 3 biological replicates of dark and L4h mRNA were applied for two-color-microarray using custom designed alternative-splicing array from Agilent Technologies.

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