Pathogenic human mitochondrial tRNA variants impair RNA processing by compromising 5' leader removal

Area of Science:

• Molecular Biology
• Genetics
• Biochemistry

Background:

• The human mitochondrial genome (mtDNA) encodes essential oxidative phosphorylation proteins and RNAs.
• Polycistronic transcripts are processed by mitochondrial RNases, with tRNAs acting as processing signals.
• Mutations in mitochondrial tRNA genes are linked to pathologies, but their processing impact is unclear.

Purpose of the Study:

• To investigate the impact of human mitochondrial tRNA sequence variations on RNase-catalyzed processing.
• To use tRNA ^Tyr as a model to study 5' leader and 3' trailer hydrolysis defects.
• To establish a framework for predicting variant effects on mitochondrial tRNA processing.

Main Methods:

• In vitro assays using human mitochondrial tRNA ^Tyr variants.
• Analysis of 5' leader and 3' trailer hydrolysis by mitochondrial RNases.
• Assessment of tRNA release in tandem tRNA clusters.

Main Results:

• Nucleotide variations near catalytic interfaces, especially in the acceptor stem, caused significant processing defects.
• These variations impaired 5' processing and prevented downstream tRNA release in tandem clusters.
• Structural position relative to processing enzymes correlates with processing efficiency.

Conclusions:

• Mitochondrial tRNA sequence variations disrupt RNase-mediated processing.
• Defects in tRNA processing contribute to mtDNA-linked pathologies.
• This study provides mechanistic insights and a predictive framework for variant effects.