The study used phospho‑proteomics to uncover rapid phosphorylation changes in Arabidopsis seedlings upon light or sucrose exposure, identifying RS41 as a hyperphosphorylated SR protein. By creating single and higher‑order mutants of four RS genes, the authors demonstrated that these RS proteins are essential for photomorphogenic development and regulate light‑dependent alternative splicing, with loss of all four causing sterility.

The study investigated how Arabidopsis thaliana SR protein kinases (AtSRPKs) regulate alternative RNA splicing by using chemical inhibitors of SRPK activity. Inhibition with SPHINX31 and SRPIN340 caused reduced root growth and loss of root hairs, accompanied by widespread changes in splicing and phosphorylation of genes linked to root development and other cellular processes. Multi‑omics analysis (transcriptomics and phosphoproteomics) revealed that AtSRPKs modulate diverse splicing factors and affect the splicing landscape of numerous pathways.

The study used TurboID-based proximity labeling coupled with mass spectrometry to map the Arabidopsis alternative splicing machinery centered on ACINUS, PININ, and SR45, identifying 298 high-confidence components and revealing that splicing is tightly linked to transcription and other RNA processing steps. Bioinformatic and genetic analyses, including O-glycosylation double mutants, demonstrated both conserved and plant‑specific regulatory networks and highlighted the role of sugar modifications in modulating splicing.

The study demonstrates that abscisic acid (ABA) accumulates in darkness to suppress cotyledon opening during seedling deetiolation, and that light exposure lifts this repression, enabling cotyledon aperture. Genome‑wide transcriptional and alternative‑splicing changes accompany this process, and the light‑dependent regulation requires the splicing factors RS40 and RS41, whose activity is repressed in the dark.