Distinct clinical phenotypes in two unrelated patients with mutations in the TRNT1 gene encoding tRNA nucleotidyl transferase. F. Sasarman1, 2, I. Thiffault2, 3, W. Weraarpachai1, S. Salomon1, C. Maftei2, J. Gauthier2, N. Webb1, 2, O. Elpeleg4, C. Brunel-Guitton2, G. Mitchell2, E. A. Shoubridge1 1) Molecular Neurogenetics, Montreal Neurological Institute and McGill University, Montreal, Quebec, Canada; 2) Medical Genetics, Pediatrics, CHU Sainte-Justine, Université de Montréal, Montreal, Quebec, Canada; 3) Center for Pediatric Genomic Medicine, Children's Mercy Hospital, Kansas City, MO, USA; 4) Genetic and Metabolic Diseases, Hadassah Medical Center, Jerusalem, Israel.

   Addition of the trinucleotide CCA to the 3end of transfer RNAs (tRNAs) is required for amino acid attachment, tRNA positioning on the ribosome and translation termination. The enzyme responsible is TRNT1 (tRNA nucleotidyl transferase), active both in the cytoplasm and mitochondria. We describe the first identification of mutations in the TRNT1 gene [MIM 612907], using exome sequencing in two unrelated patients with largely non-overlapping clinical phenotypes. Patient 1, a girl born to consanguineous parents, presented at 3 weeks of age with a crisis of lactic acidosis, evolved to severe developmental delay, hypotonia, microcephaly, seizures, progressive cortical atrophy, neurosensorial deafness, sideroblastic anemia, renal Fanconi syndrome and nephrocalcinosis and died at 21 months during a febrile episode with severe lactic acidosis and hepatic failure. Patient 2, a boy with a milder systemic phenotype, presented at 3.5 years with gait ataxia, dysarthria, gross motor regression, hypotonia, ptosis, horizontal ophthalmoplegia, and had abnormal signals in brainstem and dentate nuclei. He never had seizures or acidotic crises. Exome sequencing of both patients revealed mutations in the TRNT1 gene at evolutionary conserved positions: a homozygous c.C443T; p.A148V mutation in Patient 1, and combined heterozygous mutations c.383A>G; p.D128G and c.518A>T; p.Y173F in Patient 2. Levels of mutant TRNT1 protein in fibroblasts from Patient 1 were 10% of control, while they were normal in fibroblasts from Patient 2, suggesting a possible genotype-phenotype correlation. Muscle from Patient 1 showed a generalized decrease in the enzymatic activity of all mitochondrial respiratory chain complexes, while mitochondrial translation, respiratory chain assembly and function were normal in fibroblasts despite reduced levels of TRNT1 protein. Knockdown of TRNT1 to immunologically-undetectable levels abolished mitochondrial translation in patient fibroblasts, and had a differential effect on the steady-state levels of individual tRNAs: mitochondrial tRNASer(AGY), which has the most non-canonical structure, was undetectable, while cytoplasmic tRNALys(UUU) and tRNAMet were 30% of control. In control muscle, TRNT1 levels are 10% of the levels in fibroblasts, suggesting that muscle may be particularly vulnerable to TRNT1 mutations.

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