Progressive mitochondrial and autophagic dysfunction during RGC development driven by a LHON-associated mitochondrial tRNAThr mutation.

Leber's hereditary optic neuropathy (LHON) is a genetically inherited disease of the eye triggered by mtDNA mutations, leading to degeneration of RGCs. We previously reported that the mitochondrial tRNAThr (MT-TT) 15927G > A homoplasmic mutation disrupted the base pairing (28C-42G) conserved in the anticodon stem of tRNAThr, impairing t6A modification, aminoacylation, and steady-state tRNAThr levels, ultimately resulting in mitochondrial dysfunction. However, the absence of suitable animal and cell models for LHON has delayed efforts to elucidate disease pathophysiology, particularly tissue-specific effects. In this study, RGC-like cells were generated from iPSCs derived from a Chinese family member carrying the m.15927G > A mutation and from a control subject without this mutation. Mitochondrial dysfunction and autophagy/mitophagy defects were investigated at three differentiation stages: iPSCs, NPCs, and RGC-like cells. Both iPSCs and NPCs harboring this mutation exhibited abnormal mitochondrial dynamics, mitochondrial dysfunction, and defects in autophagy and mitophagy. RGC-like cells carrying the mutation showed significant abnormalities, including shorter neurites, imbalanced mitochondrial dynamics, elevated ROS production, reduced mitochondrial membrane potential, and impaired autophagy and mitophagy. These results indicate that the m.15927G > A mutation induces progressive mitochondrial dysfunction and developmental defects in RGCs, providing new insights into LHON pathogenesis and establishing a valuable model for future therapeutic development.