Nonsense-mediated decay of alternative pre-mRNA splicing variants is a major determinant of the Arabidopsis steady state transcriptome

The nonsense-mediated decay (NMD) surveillance pathway can recognize erroneous transcripts and physiological mRNAs, such as precursor mRNA alternative splicing (AS) variants. Currently, information on the global extent of coupled AS and NMD remains scarce and even absent for any plant species. To address this, we conducted transcriptome-wide splicing studies using Arabidopsis thaliana mutants in the NMD factor homologs UP FRAMESHIFT1 (UPF1) and UPF3 as well as wild-type samples treated with the translation inhibitor cycloheximide. Our analyses revealed that at least 17.4% of all multi-exon, protein-coding genes produce splicing variants that are targeted by NMD. Moreover, we provide evidence that UPF1 and UPF3 act in a translation-independent mRNA decay pathway. Importantly, 92.3% of the NMD-responsive mRNAs exhibit classical NMD-eliciting features, supporting their authenticity as direct targets. Genes generating NMD-sensitive AS variants function in diverse biological processes, inclu...