Identification of 25 new mutations in 40 unrelated Spanish Niemann-Pick type C patients: genotype-phenotype correlations

To better characterize Niemann-Pick type C (NPC) in Spain and improve genetic counselling, molecular analyses were carried out in 40 unrelated Spanish patients. The search identified 70/80 alleles (88%) involving 38 different NPC1 mutations, 26 of which are described for the first time. No patient with NPC2 mutations was identified. The novel NPC1 mutations include 14 amino acid substitutions [R372W (c.1114C>T), P434L (c.1301C>T), C479Y (c.1436G>A), K576R (c.1727G>A), V727F (c.2179G>T), M754K (c.2261T>A), S865L (c.2594C>T), A926T (c.2776G>A), D948H (c.2842G>C), V959E (c.2876T>A), T1036K (c.3107C>A), T1066N (c.3197C>A), N1156I (c.3467A>T) and F1224L (c.3672C>G)], four stop codon [W260X (c.780G>A), S425X (c.1274C>A), C645X (c.1935T>A) and R1059X (c.3175C>T)], two donor splice-site mutations [IVS7+1G>A (g.31432G>A) and IVS21+2insG (g.51871insG)], one in-frame mutation [N961_F966delinsS (c.2882del16bpins1bp)] and five frameshift mutations [V299fsX8 (c.895insT), A558fsX11 (c.1673insG), C778fsX10 (c.2334insT), G993fsX3 (c.2973_78delG) and F1221fsX20 (c.3662delT)]. We also identified three novel changes [V562V (c.1686G>A), A580A (c.1740C>G) and A1187A (c.3561G>T)] in three independent NPC patients and five polymorphisms that have been described previously. The combination of these polymorphisms gave rise to the establishment of different haplotypes. Linkage disequilibrium was detected between mutations C177Y and G993fsX3 and specific haplotypes, suggesting a unique origin for these mutations. In contrast, I1061T mutation showed at least two different origins. The most prevalent mutations in Spanish patients were I1061T, Q775P, C177Y and P1007A (10, 7, 7 and 5% of alleles, respectively). Our data in homozygous patients indicate that the Q775P mutation correlates with a severe infantile neurological form and the C177Y mutation with a late infantile clinical phenotype.     

Acid sphingomyelinase: identification of nine novel mutations among Italian Niemann Pick type B patients and characterization of in vivo functional in-frame start codon

Niemann Pick disease (NPD) is an autosomal recessive disorder due to the deficit of lysosomal acid sphingomyelinase, which results in intracellular accumulation of sphingomyelin. In the present work we studied 18 patients with NPD type B, including five individuals who presented an intermediate phenotype characterised by different levels of neurological involvement. We identified nine novel mutations in the SMPD1 gene including six single base changes c.2T>G, c.96G>A, c.308T>C, c.674T>C, c.732G>C, c.841G>A (p.M1_W32del, p.W32X, p.L103P, p.L225P, p.W244C, p.A281T) and three frameshift mutations c.100delC, c.565dupC, c.575dupC (p.G34fsX42, p.P189fsX1 and p.P192fsX14). The novel c.2T>G (p.M1_W32del) mutation inactivates the first in-frame translation start site of the SMPD1 gene and in the homozygous status causes NPD type B indicating that in'vivo translation of wild type SMPD1 initiates from the first in-frame ATG. Moreover, the new c.96G>A (p.W32X) introduces a premature stop codon before the second in-frame ATG. As a consequence of either c.2T>G (p.M1_W32del) or c.96G>A (p.W32X), impaired translation from the first in-frame ATG results in a mild NPD-B phenotype instead of the severe phenotype expected for a complete deficiency of the enzyme, suggesting that when the first ATG is not functional, the second initiation codon (ATG33) still produces a fairly functional sphingomyelinase. Analysis of the patients'clinical and molecular data demonstrated that all five patients with the intermediate phenotype carried at least one severe mutation. No association between the onset of pulmonary symptoms and genotype was observed. Finally, the presence of c.96G>A (p.W32X), the most frequent allele among Italian NPD type B population, and c.1799G>C (p.R600P) as compound heterozygotes in association with severe mutations suggested a beneficial effect for both mutations.     

Mutation spectrum in Taiwanese patients with phenylalanine hydroxylase deficiency and a founder effect for the R241C mutation

The spectrum of phenylalanine hydroxylase (PAH) gene mutations was determined in 25 families of hyperphenylalaninemia identified by a neonatal screening program in Taiwan. The coding sequence and exon-flanking intron sequences of PAH gene were amplified and sequenced. Mutations were identified in forty-five of the 50 chromosomes. R241C was the most common mutation (36% of the chromosomes), followed by R408Q (14% of the chromosomes). The remaining mutations were rare and seven mutations have not been reported before: p.F233L (c.697T>C), p.R252Q (c.756G>A), p.E286K (c.856G>A), p.G312V (c.935G>T), p.P314T (c.940C>A), p.I95del (c.284_286delTCA), and p.T81fsX6 (c.241_256del). Both p.R241C and p.R408Q are classified as mild phenylketonuria (PKU) or mild hyperphenylalaninemia (MHP) mutation, which may explain the fact that classical PKU is very rare in Taiwan (n=4, or one in 413,035). This strong founder effect for the p.R241C mutation has been described neither in the Caucasian populations, nor in other reports from Chinese. Since most of the populations in Taiwan are derived from Southeastern China, the spectrum of PAH gene mutations in Southeastern China should be different from other Chinese populations. This report not only disclose a specific spectrum of PAH gene mutation in Taiwan, but may also give clues to the movement of populations in Mainland China.     

Molecular analysis of mucopolysaccharidosis type VI in Poland, Belarus, Lithuania and Estonia

Mucopolysaccharidosis VI (MPS VI) is a rare autosomal recessive disorder caused by a deficiency of N-acetylgalactosamine-4-sulfatase (ARSB). Over 130 ARSB gene mutations have been identified thus far and most mutations are unique to individual families. We aimed to analyze the spectrum of mutations in the ARSB gene responsible for the disorder in Poland, Belarus and Baltic States. Twenty one families with MPS VI patients, in whom diagnosis was confirmed biochemically and enzymatically, were studied. Direct sequencing of patient genomic DNA was used to identify ARSB mutations. In total, fourteen different disease-causing mutations were found. Three novel mutations included insertion c.375_376insT, a missense mutation c.499G>A (p.G167R) and deletion/insertion c.750_754delinsCCTGAAGTCAAG. We also report 11 previously described mutations (p.A33V, p.W57C, p.Q88X, p.T92K, p.Q97X, p.R152W, p.R160Q, p.R160X, p.Y210C, p.Y266S, p.G302R). The mutation p.R152W was present at a high prevalence of 50% (21/42) the mutated alleles in this group of patients. High prevalence of p.R152W mutation in Poland, Belarus and Baltic States indicates a possible founder effect and suggests that screening for this mutation may be appropriate in MPS VI patients from this region. Our study has also provided evidence to support genotype-phenotype correlation.     

The spectrum of aldolase B (ALDOB) mutations and the prevalence of hereditary fructose intolerance in Central Europe

We investigated the molecular basis of hereditary fructose intolerance (HFI) in 80 patients from 72 families by means of a PCR-based mutation screening strategy, consisting of heteroduplex analysis, restriction enzyme digest, DNA single strand electrophoresis, and direct sequencing. For a subset of patients mutation screening with DHPLC was established which turned out to be as fast and as sensitive as the more conventional methods. Fifteen different mutations of the aldolase B (ALDOB) gene were identified in HFI patients. As in smaller previous studies, p.A150P (65%), p.A175D (11%) and p.N335K (8%) were the most common mutated alleles, followed by c.360_363delCAAA, p.R60X, p.Y204X, and c.865delC. Eight novel mutations were identified in eight families with HFI: a small indel mutation (c.1044_1049delTTCTGGinsACACT), two small deletions (c.345_372del28; c.841_842delAC), two splice site mutations (c.113-1G>A, c.799+2T>A), one nonsense mutation (c.612T>G (p.Y204X)), and two missense mutations (c.532T>C (p.C178R), c.851T>C (p.L284P)). By mutation screening for the three most common ALDOB mutations by DHPLC in 2,000 randomly selected newborns we detected 21 heterozygotes. Based on these data and after correction for less common and private ALDOB mutations, HFI prevalence in central Europe is estimated to be 1:26,100 (95% confidence interval 1: 12,600-79,000).     

Ten novel mutations in VMD2 associated with Best macular dystrophy (BMD)

Mutations in the vitelliform macular dystrophy 2 (VMD2) gene encoding besrtophin are responsible for Best macular dystrophy (BMD), a juvenile-onset autosomal dominant disorder of the central retina. Here, we report ten novel VMD2 mutations identified in clinically diagnosed BMD patients. The heterozygous alterations include nine missense mutations (c.32A>T, c.76G>C, c.85T>C, c.122T>C, c.122T>C, c.310G>C, c.722C>A, c.880C>G, c.893T>C) resulting in amino acid changes (respectively: Asn11Ile, Gly26Arg, Tyr29His, Leu41Pro, Trp102Arg, Asp104His, Thr241Asn, Leu294Val and Phe298Ser) located within four previously defined hotspot regions of the gene. In addition, a silent exonic mutation (c.624G>A) was identified in a two generation BMD pedigree. To determine a possible pathogenic effect of this variant, the consequences on splicing behaviour and potential exonic splice enhancer (ESE) motifs were analyzed. Finally, a 1-bp deletion (c.779delC) resulting in a frameshift mutation (Pro260fsX288) was found in exon 7, representing the first case of a potential frameshift mutation that affects the N-terminal half of the VMD2 protein. Besides a dominant negative effect which is likely attributable to the identified missense mutations, the deletion mutation suggests haploinsufficiency as an infrequent disease-causing mechanism in BMD.     

Identification of the molecular defects in Spanish and Argentinian mucopolysaccharidosis VI (Maroteaux-Lamy syndrome) patients, including 9 novel mutations

Maroteaux-Lamy syndrome, or mucopolysaccharidosis VI (MPS VI), is an autosomal recessive lysosomal storage disorder caused by a deficiency of N-acetylgalactosamine-4-sulfatase or arylsulfatase B (ARSB). We aimed to analyze the spectrum of mutations responsible for the disorder in Spanish and Argentinian patients, not previously studied. We identified all the ARSB mutant alleles, nine of them novel, in 12 Spanish and 4 Argentinian patients. The new changes were as follows: six missense mutations: c.245T>G [p.L82R], c.413A>G [p.Y138C], c.719C>T [p.S240F], c.922G>A [p.G308R], c.1340G>T [p.C447F] and c.1415T>C [p.L472P]; one nonsense mutation: c.966G>A [p.W322X]; and two intronic changes involving splice sites: c.1142+2T>A, in the donor splice site of intron 5, which promotes skipping of exon 5, and c.1143-1G>C, which disrupts the acceptor site of intron 5, resulting in skipping of exon 6. We also report 10 previously described mutations as well as several non-pathogenic polymorphisms. Haplotype analysis indicated a common origin for most of the mutations found more than once. Most of the patients were compound heterozygotes, whereas only four of them were homozygous. These observations confirm the broad allelic heterogeneity of the disease, with 19 different mutations in 16 patients. However, the two most frequent mutations, c.1143-1G>C and c.1143-8T>G, present in both populations, accounted for one-third of the mutant alleles in this group of patients.     

Functional assays testing pathogenicity of 14 cystathionine-beta synthase mutations

In this study, 14 CBS alleles from homocystinuric patients were expressed heterologously in E. coli and their enzyme activities were assayed in vitro. Additionally, mutant CBS proteins were visualized by Western blot from denaturing and non-denaturing polyacrylamide gels. The 14 mutations characterized were: p.R125W (c.373C>T), p.G148R (c.442G>A), p.M173V (c.517A>G), p.T191M (c.572C>T), p.A226T (c.676G>A), p.C275Y (c.824G>A), p.R336C (c.1006C>T), p.R336H (c.1007G>A), p.L338P (c.1013T>C), p.S349N (c.1046G>A), p.R379Q (c.1136G>A), p.L456P (c.1367T>C), p.G522fsX540 (c.1566delG), and p.R548Q (c.1643G>A). Eleven of the mutant alleles exhibited an activity lower than 4% of the wild-type protein. In contrast, mutations p.A226T and p.M173V presented 20% and 40% of the wild-type activity, respectively, whereas the activity of p.R548Q was up to 60% of the wild-type. This suggests that it is a new rare variant rather than a pathogenic mutation. Most of the mutated proteins exhibited a decreased signal in Western blot analyses. The non-denaturing PAGE revealed that the wild-type protein retained the capacity to form a multimeric quaternary structure, whereas in the mutations p.M173V, p.A226T, and p.G548Q, this structure grade was dramatically reduced and was completely absent in the rest of the mutations.     

Mutational analysis of mucopolysaccharidosis type VI patients undergoing a trial of enzyme replacement therapy

Mucopolysaccharidosis type VI (MPS VI), or Maroteaux-Lamy syndrome, is a lysosomal storage disorder caused by a deficiency of N-acetylgalactosamine-4-sulfatase (ARSB). Seven MPS VI patients were chosen for the initial clinical trial of enzyme replacement therapy. Direct sequencing of genomic DNA from these patients was used to identify ARSB mutations. Each individual exon of the ARSB gene was amplified by PCR and subsequently sequenced. Nine substitutions (c.289C>T [p.Q97X], c.629A>G [p.Y210C], c.707T>C [p.L236P], c.936G>T [p.W312C], c.944G>A [p.R315Q], c.962T>C [p.L321P], c.979C>T [p.R327X], c.1151G>A [p.S384N], and c.1450A>G [p.R484G]), two deletions (c.356_358delTAC [p.Y86del] and c.427delG), and one intronic mutation (c.1336+2T>G) were identified. A total of 7 out of the 12 mutations identified were novel (p.Y86del, p.Q97X, p.W312C, p.R327X, c.427delG, p.R484G, and c.1336+2T>G). Two of these novel mutations (p.Y86del and p.W312C) were expressed in Chinese hamster ovary cells and analyzed for residual ARSB activity and mutant ARSB protein. The two common polymorphisms c.1072G>A [p.V358M] and c.1126G>A [p.V376M] were identified among the patients, along with the silent mutation c.1191A>G. Cultured fibroblast ARSB mutant protein and residual activity were determined for each patient, and, together with genotype information, were used to predict the expected clinical severity of each MPS VI patient.     

Kallmann syndrome: 14 novel mutations in KAL1 and FGFR1 (KAL2)

Kallmann syndrome (KAL) combines hypogonadotropic hypogonadism and anosmia. Hypogonadism is due to Gonadotropin Releasing Hormone (GnRH) deficiency and anosmia is related to hypoplasia of the olfactory bulbs. Occasional symptoms include renal agenesis, bimanual synkinesia, cleft lip palate, dental agenesis. KAL is genetically heterogeneous and two genes have so far been identified, namely KAL1 (Xp22.3) and FGFR1/KAL2 (8p12), which underlie the X chromosome‐linked form and an autosomal dominant form of the disease, respectively. We studied a cohort of 98 unrelated Caucasian KAL patients. We identified KAL1 mutations in 14 patients, of which 7 (c.3G>A (p.M1?), g.IVS1+1G>T, c.570_571insA (p.R191fsX14), c.784G>C (p.R262P), c.958G>T (p.E320X), c.1651_1654delinsAGCT (p.P551_E552delinsSX), c.1711T>A (p.W571R)) have not been previously reported. In addition, we found FGFR1 mutations in 7 patients, namely c.303G>A (p.V102I), C.385A>C (p.D129A), c.810G>A (p.V273M), c.1093_1094delAG (p.R365fsX41), c.1561G>A (p.A520T), c.1836_1837insT (p.Y613fsX42), c.2190C>G (p.Y730X), all of which were novel mutations. In this study, unilateral renal agenesis and bimanual synkinesia were exclusively found associated with KAL1mutations, cleft palate and dental agenesia with FGFR1mutations. © 2004 Wiley‐Liss, Inc.     

Molecular analysis of Bruton’s tyrosine kinase gene in Spain

Mutations in Bruton’s tyrosine kinase (BTK) gene result in X linked agammaglobulinemia (XLA). Using Single Strand Conformation Polymorphism (SSCP) followed by direct sequencing 21 mutations were found in 27 patients with an XLA phenotype from 21 unrelated families. We identified 13 novel and 8 known mutations: seven missense (R288W, R544G, P566S, K430E; K374N, L512P, R544S), 5 nonsense (Q196X, Y361X, L249X, Q612X, Q466X), 2 deletions of one nucleotide (A207fsX216, Q612fsX648), 2 deletion‐insertions (V219fsX227, K218fsX228), one insertion of two nucleotides (S572fsX587) and 4 point mutations in donor/acceptor splice sites (g.IVS1+1G>C, g.IVS6+5G>A, g.IVS10+1G>T, g.IVS13‐1GG>CT). Carrier detection was performed in 18 mothers. Only in one case the mutation was found to be de novo. Additionally, BTK mutations were not found in four patients without family history, but with XLA‐compatible phenotype. Hum Mutat 18:84, 2001. © 2001 Wiley‐Liss, Inc.     

Novel MLH1 mutations and a novel MSH2 polymorphism identified by SSCP and DHPLC in Portuguese HNPCC families

Mismatch repair genes MSH2 and MLH1 are the two major genes implicated in hereditary nonpolyposis colorectal cancer. For the past years, we have successfully searched for mutations in both genes in affected Portuguese families, by SSCP and DNA sequencing analysis but because of the advantages that DHPLC offers, we have established conditions in our laboratory to use this new method. While screening for mutations by both methods, in 35 individuals belonging to HNPCC Portuguese families, 4 novel MLH1 mutations (c.307-1G>C; c.1023delG [p.R341fsX366]; c.2154_2155delCA [p.H718fsX721], c.2154_2155dupCA [p.I719fsX782]), an unclassified variant (c.-28A>T) and one silent MSH2 polymorphism (c.2766T>C) have been identified.     

Identification of the bruton tyrosine kinase (BTK) gene mutations in 20 Australian families with X-linked agammaglobulinemia (XLA)

X-linked agammaglobulinemia (XLA) is an immunodeficiency caused by mutations in the Bruton tyrosine kinase (BTK) gene. Twenty Australian patients with an XLA phenotype, from 15 unrelated families, were found to have 14 mutations. Five of the mutations were previously described c.83G>A (p.R28H), c.862C>T (p.R288W), c.904G>A (p.R302G), c.1535T>C (p.L512P), c.700C>T (p.Q234X), while nine novel mutations were identified: four missense c.82C>A (p.R28S), c.494G>A (p.C165Y), c.464G>A (p.C155Y), c.1750G>A (p.G584E), one deletion c.142_144delAGAAGA (p.R48_G50del), and four splice site mutations c.241-2A>G, c.839+4A>G, c.1350-2A>G, c.1566+1G>A. Carrier analysis was performed in 10 mothers and 11 female relatives. The results of this study further support the notion that molecular genetic testing represents an important tool for definitive and early diagnosis of XLA and may allow accurate carrier status and prenatal diagnosis.     

Identification of novel MLC1 mutations in Chinese patients with megalencephalic leukoencephalopathy with subcortical cysts (MLC)

Megalencephalic leukoencephalopathy with subcortical cysts (MLC) is an autosomal, recessively inherited disease caused by mutations in the MLC1 gene. Most of the previously published studies have been carried out in ethnic populations other than the Chinese. In this study, the analysis of clinical features and MLC1 mutation screening were performed in 13 Chinese patients for the first time. A total of 10 MLC1 mutations were identified in these patients, including five novel missense mutations (c.65G>A, p.R22Q; c.95C>T, p.A32V; c.218G>A, p.G73E; c.823G>A, p.A275T; c.832T>C, p.Y278H), one novel splicing mutation (c.772-1G>C in IVS9-1), one novel small deletion (c.907_930del, p.V303_L310del), one known nonsense mutation (c.593delCTCA, p.Y198X) and two known missense mutations (c.206C>T, p.S69L; c.353C>T, p.T118M). Mutation c.772-1G>C in IVS9-1, accounting for 27.3% (3/11) of the total number of genetically confirmed patients found in this study, is thus a putative hot-spot mutation in the present study group. The existence of a unique MLC1 mutation spectrum in Chinese MLC patients was shown. A systemic study to assess the mutation spectra in different populations should be undertaken.     

Clinical characteristics and molecular analysis of 21 Chinese children with congenital agammaglobulinemia

Congenital agammaglobulinemia is a humoral primary immunodeficiency and affected patients have extremely low levels of peripheral B cells and profound deficiency of all immunoglobulin isotypes. Mutations of the Bruton's tyrosine kinase (BTK) gene are responsible for most of the congenital agammaglobulinemia. In this study, the phenotypes of congenital agammaglobulinemia were investigated in 21 male children from 21 unrelated Chinese families. Sixteen different mutations of BTK gene were identified in 18 patients, and three patients did not have BTK gene mutations. Nine mutations had been reported previously including one gross deletion (c.722_2041del), one missense mutation (c.1764G>T), three non-sense mutations (c.194C>A, c.895C>T and c.1821G>A) and four invariant splice-site mutations (c.971+2T>C, c.1481+2T>A, c.1482-2A>G, c.1699-2A>G). Seven novel mutations were identified (c.373_441del, c. 504delG, c.537delC, c.851delA, c.1637G>A, c.1879T>C and c. 1482_1882 del). Ten of the eighteen mutations of BTK gene were located in the TK domain, four in the PH domain, three in the SH3 domain and one spanned the TH, SH3, SH2 and TK domain. Candidate genes of autosomal-recessive agammaglobulinemia, including IGHM, CD79a, CD79b and IGLL1, were screened in three patients without mutations in the BTK gene. A compound heterozygosity mutation in the IGHM gene (c.1956G>A, c.175_176insC) was identified in one patient. The results of our study further support that molecular genetic testing represents an important tool for early confirmed diagnosis of congenital agammaglobulinemia and may allow accurate carrier detection and prenatal diagnosis.     

Identification of nine novel arylsulfatase a (ARSA) gene mutations in patients with metachromatic leukodystrophy (MLD)

Metachromatic leukodystrophy (MLD) is a rare autosomal recessive disorder caused by mutations of the arylsulfatase A (ARSA) gene. We have investigated more than fifty MLD patients using allele-specific PCR assays to detect the pseudodeficiency (PD) allele and several common MLD mutations, followed by comprehensive nucleotide sequencing of the ARSA gene to detect rare or private mutations. Here we report the identification of nine novel microlesions in the ARSA gene: five missense mutations (c.464C>T, c.542T>A, c.916T>C, c.973G>A, c.1286A>C), three frameshift mutations (c.205_206delTG, c.489_495del, c.1483_1486dup), and one splice donor site mutation (c.973+1G>A). Comprehensive mutation detection has facilitated carrier detection and prenatal diagnosis for several at-risk MLD families.     

Functional in vitro characterization of 14 SMPD1 mutations identified in Italian patients affected by Niemann Pick Type B disease

Niemann Pick disease (NPD) is an autosomal recessive lysosomal storage disorder caused by the deficient activity of acid sphingomyelinase due to mutations in the SMPD1 gene. We functionally characterized three novel SMPD1 mutations and 11 already reported in the Italian population. Mutant alleles were studied for enzyme activity and protein processing in transiently transfected COS-1 cells. The c.96G>A, c.100delG, c.565dupC, and c.575dupC (p.W32X, p.G34fsX42, p.P189fsX1, and p.P192fs14) alleles expressed no immunoreactive protein and consequently no enzyme activity. In contrast, cells transfected with mutants c.308T>C, c.389T>C, c.674T>C, c.732G>C, c.841G>A, c.1687G>A, c.1799G>A, and c.1799G>C (p.L103P, p.V130A, p.L225P, p.W244C, p.A281T, p.D563Y, p.R600H, p.R600P) expressed protein levels comparable to wild-type ASM expressing cells. Only three of these constructs, c.389T>C, c.1687G>A, and c.1799G>A (p.V130A, p.D563Y, p.R600H), retained residual activity while the other five expressed very low or no enzyme activity. As expected, the c.1669underscore;1670delGT (p.V557fsX18) mutant expressed a completely inactive truncated protein. Interestingly, the c.2T>G (p.M1_W32del) mutant expressed 26.9% of the wild type activity, even though no ASM protein was detected by Western blot analysis, suggesting that the amount of produced enzyme is below detection levels. The results presented in this study are consistent with the wide phenotype variability found in NP type B patients and provide valuable insights into the molecular basis of the disease.     

Identification of mutations in the GNPTA (MGC4170) gene coding for GlcNAc-phosphotransferase alpha/beta subunits in Korean patients with mucolipidosis type II or type IIIA

Mucolipidosis types II and III are autosomal recessive inherited diseases caused by a deficiency in the lysosomal enzyme N-acetylglucosamine-1 phosphotransferase (GlcNAc-phosphotransferase), which adds phosphate to function as a recognition marker for the uptake and transport of lysosomal enzymes. We investigated mutations in the GNPTA (MGC4170) gene, which codes for the alpha/beta subunits of phosphotransferase, and in the GNPTAG gene, which codes for its gamma subunits in five Korean patients with mucolipidosis type II or IIIA. We identified seven mutations in the GNPTA gene, but none in GNPTAG. The mutations in type II patients included p.Q104X (c.310C>T), p.R1189X (c.3565C>T), p.S1058X (c.3173C>G), p.W894X (c.2681G>A), and p.H1158fsX15 (c.3474_3475delTA), all of which are nonsense or frameshift mutations. However, a splicing site mutation, IVS13+1G>A (c.2715+1G>A) was detected along with a nonsense or a frameshift mutation (p.R1189X or p.E858fsX3 (c.2574_2575delGA)) in two mucolipidosis type IIIA patients. This report shows that mutations in the GNPTA gene coding for the alpha/beta subunits of phosphotransferase, and not mutations in the GNPTAG gene, account for most of the genetic mutations found in Korean patients with mucolipidosis type II or IIIA.     

Novel GNE mutations in Italian families with autosomal recessive hereditary inclusion-body myopathy

The most common form of autosomal recessive (AR) hereditary inclusion-body myopathy (HIBM), originally described in Persian-Jewish families, is characterized by onset in early adult life with weakness and atrophy of distal lower limb muscles, which progress proximally and relatively spare the quadriceps. AR HIBM is associated with mutations in the UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase gene (GNE) on chromosome 9p12-13. In the present study we have identified seven novel GNE mutations in patients from five unrelated Italian families with clinical and pathologic features indicative of AR HIBM. Four were missense mutations (c.1556A>G [p.N519S], c.79C>T [p.P27S], c.1798G>A [p.A600T] and c.616G>A [p.G206S]), two consisted in a single-base deletion (c.616delG [p.G206fsX4] and c.1130delT [p.I377fsX16]) and one in an intronic single-base insertion (c.1070+2dupT). These latter findings further extend the type of GNE mutations associated with HIBM. Furthermore, in one patient we also identified the c.737G>A [p.R246Q] missense mutation that corresponds to the one previously reported in a family from the Bahamas. Interestingly, in two of our families distinct mutations affected nucleotide c.616 in exon 3 (c.616delG and c.616G>A). The possibility of specific portions of the gene being more prone to mutations remains to be elucidated.     

Five novel inactivating mutations in the thyroid peroxidase gene responsible for congenital goiter and iodide organification defect

Thyroid peroxidase (TPO) defects, typically transmitted as autosomal recessive traits, result in hypothyroid goiters with failure to convert iodide into organic iodine. We analyzed the TPO gene in 14 unrelated patients with clinical evidence of iodide organification defects. Seven of the affected individuals harbored mutations in the TPO gene; one was compound heterozygous, the others were simply heterozygous for TPO mutations. Five novel mutations have been identified, one of which was found to be a single nucleotide deletion, while the other four were single nucleotide substitutions. A frameshift mutation c.387delC was detected in exon 5 which leads to an early termination signal in exon 7 (p.N129fsX208). Two missense mutations were identified in exon 8. The first, a c.920A>C transversion that results in a p.N307T substitution, was found in two patients. The second, a c.1297G>A transition, results in p.V433M. A c.1496C>T transition was detected in exon 9 that caused the substitution p.P499L. Finally, in exon 14 a c.2422T>C transition was identified, causing a p.C808R change. In addition, the previously reported GGCC duplication in exon 8 (c.1186_1187insGGCC; p.R396fsX472) was also detected in two affected individuals, one of whom was a compound heterozygous (p.R396fsX472/p.V433M).