From the Neurogenetics Unit (A.R.B.d.P., F.F., B.D.R.d.A., I.B., C.L., D.d.C.d.S., F.K.), Neurology Department, Hospital das Clínicas da Universidade de São Paulo, Brazil; Department of Molecular Neuroscience (D.S.L., H.H.), UCL Institute of Neurology, London, UK; Leonard Wolfson Experimental Neurology Centre (D.S.L., H.H.), UCL Institute of Neurology, London, UK; Human Genome and Stem Cell Research Center (U.S.M., L.I.M.-S., F.K.), Department of Genetics and Evolutionary Biology, Instituto de Biociências, Universidade de São Paulo, Brazil; and Neuroradiology Section (L.T.L.), Hospital das Clínicas da Universidade de São Paulo, Brazil.
Mutations in PUS3, which encodes a highly conserved enzyme responsible for posttranscriptional modification of tRNA, have been shown in a single family to be a cause of nonsyndromic intellectual disability (ID).1 In this study, we used whole-exome sequencing (WES) to identify biallelic mutations in PUS3 associated with syndromic ID with dysmorphic features, white matter disease (WMD), and renal abnormalities in a nonconsanguineous family from Brazil.
We evaluated 2 sisters (figure, A) who had ID, renal abnormalities, diffuse WMD, and dysmorphic features. Their brother was similarly affected and died at age 22 years of complications of renal disease. The parents were nonconsanguineous from Northeast Brazil and Southern Italy. We obtained approval from the institutional ethics committee and written informed consent from family members.
(A) Family pedigree: 2 affected individuals were compound heterozygotes for p.Arg166Gln and p.Leu366Pro PUS3 mutations, whereas their healthy mother and sister were heterozygotes for only 1 of the mutations. (B) Sanger sequencing electropherograms showing both c.497G>A (p.Arg166Gln) and c.1097T>C (p.Leu366Pro) PUS3 mutations. (C) Vertebrate protein sequencing alignment showing conserved amino acid sites for both PUS3 mutations. (D) Scattered FLAIR hyperintensities in the cerebral white matter in P1. (E) Symmetric diffuse FLAIR hyperintensity in the cerebral white matter in P2. (F) Axial T2-weighted image discloses not only the white matter changes in P2 but also mild T2 hyperintensity in the globus pallidus bilaterally (arrows). One can notice also in P2 symmetric FLAIR hyperintensities in the cerebellar white matter (arrows in G), together with signs of brain parenchyma volume loss (E–G).
The first patient (P1; III-3; figure, A) was noted to have global developmental delay after age 1 year. As a child, she could understand short sentences, but expressive language was limited to monosyllables. She was diagnosed with profound ID but was independent in basic activities of daily living (ADLs). She had 2 focal dyscognitive seizures with secondary generalization, at age 18 and 23 years, responsive to carbamazepine. At age 37 years, asymptomatic proteinuria and reduced creatinine clearance were detected. At 44 years, she developed psychosis, with prominent auditory and visual hallucinations and episodes of aggression. Physical examination revealed her to be on the 10th centile for height, weight, and head circumference and to have genu valgum. In addition, neurologic examination revealed dysarthria, pseudobulbar affect, with impaired gait and balance.
The second patient (P2; III-6; figure, A) also presented with short stature, neurodevelopmental delay, and dysmorphic features. She was diagnosed with acute nephritis at age 6 months and coeliac disease at age 5 years. She also developed generalized phenobarbital responsive seizures. She was able to walk and could complete ADLs with assistance. At age 20 years, she developed nephrotic syndrome due to focal segmental glomerulosclerosis (FSGS) with mesangial granular deposits and positive IgM and C3 immunofluorescence. She did not respond to steroids and developed end-stage renal failure requiring hemodialysis. Physical examination revealed that she was on the 3rd centile for height, weight, and head circumference and to have gray sclera, genu valgum, pseudobulbar affect, and dysarthria. She died at age 40 years of complications of community-acquired pneumonia.
MRI identified white matter abnormalities in both patients. In P1, there were scattered T2/fluid-attenuated inversion recovery (FLAIR) hyperintensities in the cerebral white matter, more prominent in periventricular zones (figure, D). In P2, there was symmetric diffuse T2/FLAIR hyperintensity in the cerebral and cerebellar white matter with extension to the subcortical areas, mild T2 hyperintensity in the globus pallidus bilaterally, and signs of brain parenchymal volume loss (figure, E–G). There was no gadolinium enhancement or areas of restricted diffusion.
We performed WES on both patients and filtered the data for a presumed autosomal recessive inheritance pattern. There were no regions of homozygosity and no shared rare homozygous variants. However, when filtered for damaging, rare compound heterozygous mutations, we identified biallelic mutations only in PUS3 (p.Arg166Gln;p.Leu366Pro) (figure, B). Both are very rare and highly conserved residues among vertebrates. We confirmed the mutations by Sanger sequencing and demonstrated that they were inherited in trans.
The original report of PUS3-related disorder describes a family with 3 affected individuals (age: 3–15 years) from Saudi Arabia with severe ID, multifocal white matter changes, and in a single patient, epilepsy. No nephropathy was reported. WES detected a homozygous truncating mutation (p.Arg435Ter), predicting to lead to a loss of function. In our patients, affected individuals have a milder intellectual compromise and developed proteinuria/nephrotic syndrome after age 20 years. In addition, the white matter abnormalities were more extensive in our patients.
Of interest, a well-known mechanism of WMD is disruption of protein translation as is seen in mutations in mitochondrial aspartate (DARS2) or glutamate (EARS2) transfer-RNA synthase. It is also noteworthy that leukoencephalopathies usually cause progressive neurologic deterioration and not static encephalopathy with ID, as occurred in our patients.
The late-onset renal involvement observed in our patients expands the phenotype of PUS3-related disorder. Steroid-resistant FSGS has been rarely associated with mutations in mitochondrial tRNA in patients with other features typical of mitochondrial dysfunction.4,–,7 Recently, tubulopathy was reported in deficiency of tRNA N6-adenosine threonylcarbamoyltransferase, an enzyme involved in posttranscriptional tRNA modification.
This study expands the known phenotypic and allelic spectrum of PUS3-related disorder. In addition, it provides further evidence of the relevance of posttranscriptional modification of tRNA in neurologic diseases. Given these additional findings, we believe the phenotypic spectrum of PUS3 mutations to be wider than the initial report.
A.R.B. de Paiva, D.S. Lynch, and U.S. Melo report no disclosures. L.T. Lucato has received speaker honoraria from Bracco Imaging do Brasil and has served on the editorial board of Arquivos de Neuro-Psiquiatria. F. Freua, B.D.R. de Assis, I. Barcelos, C. Listik, D. Castro Santos, and L.I. Macedo-Souza report no disclosures. H. Houlden has received governmental and/or foundation/society support from the Medical Research Council (MRC) UK, the BRT, the MDA USA, Muscular Dystrophy UK, Ataxia UK, Muscular Dystrophy UK, Rosetrees Trust, the Wellcome Trust, and the National Institute for Health Research (NIHR) UCL/UCLH BRC. F. Kok has served on the editorial board of Arquivos de Neuro-Psiquiatria; holds a patent for methylmalonic acid determination by tandem mass spectrometry using stable isotope; is employed as Medical Director for Mendelics Genomic Analysis; has acted as a speaker for Actelion Pharmaceuticals at the NPC symposia and for BioMarin Pharmaceuticals at the CLN2 symposia; and has been a shareholder of Mendelics Genomic Analysis. Full disclosure form information provided by the authors is available with the full text of this article at Neurology.org/NG.
The authors thank the patients and their family for participating in this study.
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