Unraveling branch point-driven SR45a splicing dynamics in heat stress for plant adaptation.

Abstract

Heat stress impairs plant development by disrupting essential molecular processes. Alternative splicing (AS) is emerging as a key regulatory mechanism in heat responses, yet how stress-responsive AS is fine-tuned within core splicing regulators remains underexplored. Here, we uncover a heat-activated, autoregulatory splicing switch in the serine/arginine-rich factor SR45a, critically dependent on intron 4. Using an intron 4-based luciferase reporter, we show that heat enhances intron removal, promoting the full-length isoform over truncated variants. This switch relies on branch point (BP) recognition, with SR45a and cap-binding protein 20 (CBP20) coordinating BP-dependent splicing under stress. Overexpression of either protein enhances thermotolerance by stabilizing AS dynamics. HSFA2 directly activates <i>SR45a</i> transcription, linking transcriptional and splicing control. Notably, intron 4-mediated splicing is conserved in both maize and wheat, suggesting its potential as a portable regulatory module in crops. Our study establishes a stress-inducible SR45a-CBP20-BP axis and introduces an intron-based strategy for engineering thermotolerance via precision splicing control.