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

Barcelos, I

de Assis, BDR

de Castro Dos Santos, D

de Paiva, ARB

Freua, F

Houlden, H

Kok, F

Listik, C

Lucato, LT

Lynch, DS

Macedo-Souza, LI

Melo, US

UCL Discovery

CLINICAL/SCIENTIFIC NOTES OPEN ACCESSPUS3 mutations are associated with intellectualdisability, leukoencephalopathy, andnephropathyAnderson Rodrigues Brandão de Paiva, MD, David S. Lynch, MD, PhD, Uira´ Souto Melo, PhD,Leandro Tavares Lucato, MD, PhD, Fernando Freua,MD, BrunoDella Ripa de Assis, MD, Isabella Barcelos, MD,Clarice Listik, MD, Diego de Castro dos Santos, MD, Lu´cia Ineˆs Macedo-Souza, PhD, Henry Houlden, MD, PhD,and Fernando Kok, MD, PhDNeurol Genet 2019;5:e306. doi:10.1212/NXG.0000000000000306CorrespondenceDr. de Paivaarbrandaopaiva@gmail.comMutations in PUS3, which encodes a highly conserved enzyme responsible for post-transcriptional modiﬁcation of tRNA, have been shown in a single family to be a cause ofnonsyndromic intellectual disability (ID).1 In this study, we used whole-exome sequencing(WES) to identify biallelic mutations in PUS3 associated with syndromic ID with dysmorphicfeatures, white matter disease (WMD), and renal abnormalities in a nonconsanguineous familyfrom Brazil.Clinical ﬁndingsWe evaluated 2 sisters (ﬁgure, A) who had ID, renal abnormalities, diﬀuse WMD, and dys-morphic features. Their brother was similarly aﬀected and died at age 22 years of complicationsof renal disease. The parents were nonconsanguineous from Northeast Brazil and SouthernItaly. We obtained approval from the institutional ethics committee and written informedconsent from family members.The ﬁrst patient (P1; III-3; ﬁgure, A) was noted to have global developmental delay after age1 year. As a child, she could understand short sentences, but expressive language was limitedto monosyllables. She was diagnosed with profound ID but was independent in basic activitiesof daily living (ADLs). She had 2 focal dyscognitive seizures with secondary generalization, atage 18 and 23 years, responsive to carbamazepine. At age 37 years, asymptomatic proteinuriaand reduced creatinine clearance were detected. At 44 years, she developed psychosis, withprominent auditory and visual hallucinations and episodes of aggression. Physical examina-tion revealed her to be on the 10th centile for height, weight, and head circumference and tohave genu valgum. In addition, neurologic examination revealed dysarthria, pseudobulbaraﬀect, with impaired gait and balance.The second patient (P2; III-6; ﬁgure, A) also presented with short stature, neurodevelopmentaldelay, and dysmorphic features. She was diagnosed with acute nephritis at age 6 months andcoeliac 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 developednephrotic syndrome due to focal segmental glomerulosclerosis (FSGS) with mesangial granulardeposits and positive IgM and C3 immunoﬂuorescence. She did not respond to steroidsand developed end-stage renal failure requiring hemodialysis. Physical examination revealedthat she was on the 3rd centile for height, weight, and head circumference and to have grayFrom 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 Cl´ınicas da Universidade de São Paulo, Brazil; Department ofMolecular Neuroscience (D.S.L., H.H.), UCL Institute of Neurology, London, UK; LeonardWolfson 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 Biocieˆncias, Universidade de São Paulo, Brazil;and Neuroradiology Section (L.T.L.), Hospital das Cl´ınicas da Universidade de São Paulo, Brazil.Funding information and disclosures are provided at the end of the article. Full disclosure form information provided by the authors is available with the full text of this article atNeurology.org/NG.The Article Processing charge was funded by the authors.This is an open access article distributed under the terms of the Creative Commons Attribution-NonCommercial-NoDerivatives License 4.0 (CC BY-NC-ND), which permits downloadingand sharing the work provided it is properly cited. The work cannot be changed in any way or used commercially without permission from the journal.Copyright © 2019 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the American Academy of Neurology. 1sclera, genu valgum, pseudobulbar aﬀect, and dysarthria. Shedied at age 40 years of complications of community-acquiredpneumonia.Neuroimaging findingsMRI identiﬁed white matter abnormalities in both patients. InP1, there were scattered T2/ﬂuid-attenuated inversion recovery(FLAIR) hyperintensities in the cerebral white matter, moreprominent in periventricular zones (ﬁgure, D). In P2, there wassymmetric diﬀuse T2/FLAIR hyperintensity in the cerebral andcerebellar white matter with extension to the subcortical areas,mild T2 hyperintensity in the globus pallidus bilaterally, andsigns of brain parenchymal volume loss (ﬁgure, E–G). Therewas no gadolinium enhancement or areas of restricted diﬀusion.Genetic findingsWe performedWES on both patients and ﬁltered the data fora presumed autosomal recessive inheritance pattern. Therewere no regions of homozygosity and no shared rare ho-mozygous variants. However, when ﬁltered for damaging,rare compound heterozygous mutations, we identiﬁed bial-lelic mutations only in PUS3 (p.Arg166Gln;p.Leu366Pro)Figure Genetic and radiologic findings(A) Family pedigree: 2 affected individuals were compound heterozygotes for p.Arg166Gln and p.Leu366Pro PUS3 mutations, whereas their healthymother 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 PUS3mutations. (D) Scattered FLAIR hyperintensities in the cerebral white matter in P1. (E) Symmetric diffuse FLAIR hyperintensity in the cerebral whitematter in P2. (F) Axial T2-weighted image discloses not only the white matter changes in P2 but also mild T2 hyperintensity in the globus pallidusbilaterally (arrows). One can notice also in P2 symmetric FLAIR hyperintensities in the cerebellar white matter (arrows in G), together with signs of brainparenchyma volume loss (E–G).2 Neurology: Genetics | Volume 5, Number 1 | February 2019 Neurology.org/NG(ﬁgure, B). Both are very rare and highly conserved residuesamong vertebrates. We conﬁrmed the mutations by Sangersequencing and demonstrated that they were inherited intrans.DiscussionPUS3 codes for pseudourydilate synthase, a highly conservedenzyme responsible for posttranscriptional modiﬁcation oftRNA,2 which is critical for its structure, function, and sta-bility. Most of these modiﬁcations have been recently recog-nized in a growing number of human genes leading toID–FTSJ1, TRMT1, NSUN2, WDR4, ADAT3, and PUS3.3The original report of PUS3-related disorder describes a familywith 3 aﬀected individuals (age: 3–15 years) from Saudi Arabiawith severe ID, multifocal white matter changes, and in a singlepatient, epilepsy. No nephropathy was reported. WES detecteda homozygous truncatingmutation (p.Arg435Ter), predicting tolead to a loss of function. In our patients, aﬀected individuals havea milder intellectual compromise and developed proteinuria/nephrotic syndrome after age 20 years. In addition, the whitematter abnormalities were more extensive in our patients.Of interest, a well-known mechanism of WMD is disruptionof protein translation as is seen in mutations in mitochondrialaspartate (DARS2) or glutamate (EARS2) transfer-RNAsynthase. It is also noteworthy that leukoencephalopathiesusually cause progressive neurologic deterioration and notstatic encephalopathy with ID, as occurred in our patients.The late-onset renal involvement observed in our patientsexpands the phenotype of PUS3-related disorder. Steroid-resistant FSGS has been rarely associated with mutations inmitochondrial tRNA in patients with other features typical ofmitochondrial dysfunction.4–7 Recently, tubulopathy wasreported in deﬁciency of tRNA N6-adenosine threonylcarba-moyltransferase, an enzyme involved in posttranscriptionaltRNA modiﬁcation.This study expands the known phenotypic and allelic spec-trum of PUS3-related disorder. In addition, it provides furtherevidence of the relevance of posttranscriptional modiﬁcationof tRNA in neurologic diseases. Given these additional ﬁnd-ings, we believe the phenotypic spectrum of PUS3 mutationsto be wider than the initial report.AcknowledgmentThe authors thank the patients and their family for participatingin this study.Study fundingNo targeted funding reported.DisclosureA.R.B. de Paiva, D.S. Lynch, andU.S.Melo report no disclosures.L.T. Lucato has received speaker honoraria fromBracco ImagingAppendix 1. Author contributionsName Location Role ContributionAndersonRodriguesBrandão dePaiva, MDNeurogenetics Unit,Neurology Department,Hospital das Cl´ınicas daUniversidade de SãoPaulo, São Paulo, BrazilAuthor Study conceptand design,acquisition ofdata, and writingthe manuscript.David S.Lynch, MD,PhDUCL Institute ofNeurology, London,United KingdomAuthor Study conceptand design,acquisition ofdata, and writingthe manuscript.Uira´ SoutoMelo, PhDHuman Genome andStem Cell ResearchCenter, Department ofGenetics andEvolutionary Biology,Instituto de Biocieˆncias,Universidade de SãoPaulo, São Paulo, BrazilAuthor Acquisition ofdata and criticalrevision of themanuscript.LeandroTavaresLucato, MD,PhDNeuroradiologySection, Hospital dasCl´ınicas daUniversidade de SãoPaulo, São Paulo, BrazilAuthor Acquisition ofdata and criticalrevision of themanuscript.FernandoFreua, MDNeurogenetics Unit,Neurology Department,Hospital das Cl´ınicas daUniversidade de SãoPaulo, São Paulo, BrazilAuthor Acquisition ofdata and criticalrevision of themanuscript.Bruno DellaRipa de Assis,MDNeurogenetics Unit,Neurology Department,Hospital das Cl´ınicas daUniversidade de SãoPaulo, São Paulo, BrazilAuthor Acquisition ofdata and criticalrevision of themanuscript.IsabellaBarcelos, MDNeurogenetics Unit,Neurology Department,Hospital das Cl´ınicas daUniversidade de SãoPaulo, São Paulo, BrazilAuthor Acquisition ofdata and criticalrevision of themanuscript.Clarice Listik,MDNeurogenetics Unit,Neurology Department,Hospital das Cl´ınicas daUniversidade de SãoPaulo, São Paulo, BrazilAuthor Acquisition ofdata and criticalrevision of themanuscript.Diego deCastro dosSantos, MDNeurogenetics Unit,Neurology Department,Hospital das Cl´ınicas daUniversidade de SãoPaulo, São Paulo, BrazilAuthor Acquisition ofdata and criticalrevision of themanuscript.Lu´cia IneˆsMacedo-Souza, PhDHuman Genome andStem Cell ResearchCenter, Departmentof Genetics andEvolutionary Biology,Instituto de Biocieˆncias,Universidade de SãoPaulo, São Paulo, BrazilAuthor Acquisition ofdata and criticalrevision of themanuscript.HenryHoulden, MD,PhDUCL Institute ofNeurology, London,United KingdomAuthor Critical revision ofthe manuscriptFernandoKok, MD, PhDNeurogenetics Unit,Neurology Department,Hospital das Cl´ınicasda Universidade deSão Paulo, São Paulo,BrazilAuthor Critical revision ofthe manuscriptNeurology.org/NG Neurology: Genetics | Volume 5, Number 1 | February 2019 3do Brasil and has served on the editorial board of Arquivos deNeuro-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, theWellcome Trust, and the National Institute for Health Re-search (NIHR) UCL/UCLH BRC. F. Kok has served on theeditorial board of Arquivos de Neuro-Psiquiatria; holds apatent for methylmalonic acid determination by tandemmassspectrometry using stable isotope; is employed as MedicalDirector for Mendelics Genomic Analysis; has acted asa speaker for Actelion Pharmaceuticals at the NPC symposiaand for BioMarin Pharmaceuticals at the CLN2 symposia;and has been a shareholder of Mendelics Genomic Analysis.Full disclosure form information provided by the authors isavailable with the full text of this article at Neurology.org/NG.Publication historyReceived byNeurology: Genetics August 13, 2018. Accepted in ﬁnal formNovember 8, 2018.References1. Shaheen R, Han L, Faqeih E, et al. A homozygous truncating mutation in PUS3 expandsthe role of tRNA modiﬁcation in normal cognition. Hum Genet 2016;135:707–713.2. Hur S, Stroud RM. How U38, 39, and 40 of many tRNAs become the targets forpseudouridylation by TruA. Mol Cell 2007;26:189–203.3. Torres AG, Batlle E, Ribas de Pouplana L. Role of tRNA modiﬁcations in humandiseases. Trends Mol Med 2014;20:306–314.4. Dinour D, Mini S, Polak-Charcon S, Lotan D, Holtzman EJ. Progressive ne-phropathy associated with mitochondrial tRNA gene mutation. Clin Nephron2004;62:149–154.5. Scaglia F1, Vogel H, Hawkins EP, Vladutiu GD, Liu LL,Wong LJ. Novel homoplasmicmutation in the mitochondrial tRNATyr gene associated with atypical mitochondrialcytopathy presenting with focal segmental glomerulosclerosis. Am J Med Genet A2003;123A:172–178.6. Gue´ry B, Choukroun G, Noe¨l LH, et al. The spectrum of systemic involvement inadults presenting with renal lesion and mitochondrial tRNA(Leu) gene mutation.J Am Soc Nephrol 2003;14:2099–2108.7. Hotta O, Inoue CN, Miyabayashi S, Furuta T, Takeuchi A, Taguma Y. Clinical andpathologic features of focal segmental glomerulosclerosis with mitochondrialtRNALeu(UUR) gene mutation. Kidney Int 2001;59:1236–1243.4 Neurology: Genetics | Volume 5, Number 1 | February 2019 Neurology.org/NGDOI 10.1212/NXG.00000000000003062019;5; Neurol Genet Anderson Rodrigues Brandão de Paiva, David S. Lynch, Uirá Souto Melo, et al. nephropathy mutations are associated with intellectual disability, leukoencephalopathy, andPUS3This information is current as of January 16, 2019ServicesUpdated Information & http://ng.neurology.org/content/5/1/e306.full.htmlincluding high resolution figures, can be found at:References http://ng.neurology.org/content/5/1/e306.full.html##ref-list-1This article cites 7 articles, 1 of which you can access for free at: Subspecialty Collections http://ng.neurology.org//cgi/collection/mental_retardationMental retardation http://ng.neurology.org//cgi/collection/leukodystrophiesLeukodystrophies http://ng.neurology.org//cgi/collection/all_geneticsAll Geneticsfollowing collection(s): This article, along with others on similar topics, appears in the  Permissions & Licensing http://ng.neurology.org/misc/about.xhtml#permissionsits entirety can be found online at:Information about reproducing this article in parts (figures,tables) or in  Reprints http://ng.neurology.org/misc/addir.xhtml#reprintsusInformation about ordering reprints can be found online:reserved. 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PUS3 mutations are associated with intellectual disability, leukoencephalopathy, and nephropathy

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PUS3 mutations are associated with intellectual disability, leukoencephalopathy, and nephropathy

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