PCIS1 is an essential factor in mitochondrial RNA splicing and complex I biogenesis, with distinct effects in null and downregulated mutants.

PCIS1 is indispensable for plant mitochondria NAD4 and NAD7 gene expression. Null mutants abolish CI assembly and arrest embryogenesis, whereas knockouts retain partial splicing, revealing a genetic functional mt-RNA metabolism threshold. Plant mitochondrial biogenesis relies on the splicing of numerous group II introns, primarily in genes encoding complex I (CI) subunits, which require multiple nuclear-encoded cofactors. Recently, we characterized PPR co-expressed intron splicing-1 (PCIS1), an essential factor identified in silico, based on its co-expression with several pentatricopeptide repeat (PPR) proteins. Knockout of Arabidopsis PCIS1 results in embryonic arrest, but mutant lines can be maintained via embryo-specific ABI3-driven expression. The partially complemented pABI3::PCIS1 plants show growth and developmental defect phenotypes, associated with defective processing of nad2, nad4, and nad7 mitochondrial transcripts. To eliminate the potential leakiness of tissue-specific promoters and better define PCIS1 function, we analyzed the RNA and protein profiles in homozygous pcis1 seedlings generated by a modified embryo-rescue approach. Our data confirm that PCIS1 is essential for respiratory biogenesis via splicing of nad4 intron 3 and nad7 introns 1 and 2, and that null and downregulated pcis1 lines display distinct phenotypic and molecular defects. Analysis of different pcis1 mutant lines links the embryogenesis defects to impaired CI assembly and respiratory dysfunction, and reveals distinct consequences of PCIS1 knockdown versus null mutations for mt-RNA processing, OXPHOS biogenesis and plant physiology.