PDD-mediated mitochondrial and chloroplast tRNA modifications regulate cellular protein synthesis by shaping the...

Stable tRNA modifications regulate developmental processes by ensuring efficient protein synthesis across different organisms. However, our understanding of tRNA-modification enzymes in plant organelles remains relatively limited. Previously, natural variants of rice PDD (PLEIOTROPIC DEVELOPMENTAL DEFECTS) were shown to disrupt chloroplast tRNA mnm⁵s²U modification and cause growth defects. Here, we identified three critical residues (positions 145, 191, and 376) in PDD^OL essential for PDD function. We found that the pleiotropic developmental defects in the NIL-PDD^OL were associated with abnormal mitochondrial development, indicating that PDD also plays an essential role in mitochondria. Biochemical analyses confirmed that PDD dysfunction significantly reduced mitochondrial complex enzyme activities and substantially decreased mitochondrial protein accumulation. Ultra-high-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) revealed that PDD regulates τm⁵U, τm⁵s²U and mnm⁵s²U modifications in both mitochondrial and chloroplast tRNAs. RNA-seq and Ribo-seq analyses revealed that these modification deficiencies caused codon-specific translation stalling in mitochondrial and chloroplast genes, impairing translation efficiency and reducing protein levels. Additionally, the transcription and translation of numerous nuclear genes are altered in NIL-PDD^OL via retrograde regulation, with a significant up-regulation of genes involved in cytoplasmic mRNA transcription and translation processes. Our work demonstrates that a dual-targeting organelle tRNA modification enzyme regulates cellular protein synthesis by orchestrating translation of organelle genomes and nuclear genomes. These findings provide a foundation for studying translational control in organelles and highlight the functional integration of organelle activity with plant growth and development.