The mitochondrial PPR protein AS3 interacts with NPPR2 to facilitate trans-splicing of nad1 intron 1 and ensure...

In higher plants, many mitochondrial genes contain group II introns that undergo intricate splicing processes to produce mature transcripts, which are subsequently translated into functional proteins. However, the identities and specific roles of proteins involved in mitochondrial intron splicing in maize remain largely unclear. In this study, we identified a maize mutant, abnormal seed 3 (as3), through ethyl methanesulfonate (EMS) mutagenesis and mapped the causal gene using bulked segregant analysis. AS3 encodes a mitochondrion-localized P-subgroup pentatricopeptide repeat (PPR) protein containing 21 PPR motifs. Mutation of AS3 substantially reduces the trans-splicing of mitochondrial nad1 intron 1, resulting in the accumulation of nad1 exon 1 precursors and a pronounced reduction in mature nad1 transcripts. These splicing defects compromise the assembly and activity of respiratory complex I, alter mitochondrial morphology, and activate the alternative oxidase pathway, as evidenced by elevated transcript levels of alternative oxidases. Furthermore, AS3 physically interacts with the PPR protein NPPR2, whose loss of function similarly disrupts nad1 intron 1 splicing and further decreases nad1 transcript and protein abundance. Collectively, these results demonstrate that AS3 and NPPR2 physically interact and act cooperatively in the trans-splicing of nad1 intron 1, a process essential for mitochondrial function and proper kernel development in maize.