Datasets:

cnachteg

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duvel

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Interestingly, one FALS proband carried 3 variants, each of which has previously been reported as pathogenic: @GENE$ p.G38R, ANG p.P136L, and DCTN1 p.T1249I. Nine apparently sporadic subjects had variants in multiple genes (Table 4), but only two were well-established ALS mutations: @GENE$ p.G287S was found in combination with VAPB p.M170I while a subject with juvenile-onset ALS carried a de novo FUS @VARIANT$ mutation with a paternally-inherited intermediate-sized CAG expansion in ATXN2. Two SALS patients carried multiple ALS-associated variants that are rare in population databases (ANG p.K41I with VAPB p.M170I and TAF15 p.R408C with SETX p.I2547T and SETX @VARIANT$).

SOD1;392

TARDBP;7221

p.P525L;tmVar:p|SUB|P|525|L;HGVS:p.P525L;VariantGroup:62;CorrespondingGene:2521;RS#:886041390;CA#:10603390

p.T14I;tmVar:p|SUB|T|14|I;HGVS:p.T14I;VariantGroup:28;CorrespondingGene:4094;RS#:1219381953

0no label

To investigate the role of @GENE$ variations along with @GENE$ mutations for a possible combinatory allelic disease inheritance, we have screened patients with heterozygous GJB2 mutations for variants in Cx31 by sequencing. Analysis of the entire coding region of the Cx31 gene revealed the presence of two different missense mutations (N166S and A194T) occurring in compound heterozygosity along with the 235delC and 299delAT of GJB2 in 3 simplex families (235delC/@VARIANT$, @VARIANT$/A194T and 299delAT/A194T).

GJB3;7338

GJB2;2975

N166S;tmVar:p|SUB|N|166|S;HGVS:p.N166S;VariantGroup:0;CorrespondingGene:2707;RS#:121908851;CA#:118311

235delC;tmVar:c|DEL|235|C;HGVS:c.235delC;VariantGroup:1;CorrespondingGene:2706;RS#:80338943

0no label

The study revealed @GENE$ gene mutations in a majority of our cohort (33%), in accordance with the percentages already reported in the literature. Interestingly, we found just one patient with variants in BBS1, the most frequently detected gene in BBS patients. We identified a novel variant in BBS1 patient #10 c.1285dup (@VARIANT$) defined as pathogenic that segregates with phenotype together with c.46A > T (p.(Ser16Cys), defined as likely pathogenic. A new pathogenic variant in BBS2 affecting a conserved residue in the functional domain of BBsome protein (c.1062C > G; @VARIANT$) was found in compound heterozygous state in patient #1 together with the known pathogenic variant p.(Arg339*). A new homozygous nucleotide change in BBS7 that leads to a stop codon in position 255, c.763A > T, was identified in patient #3. BBS1, BBS2 and BBS7 share a partially overlapping portion of a functional domain, mutation of which results in the same disease phenotype. New pathogenic variants of @GENE$ and BBS7 lie in this portion.

BBS10;49781

BBS2;12122

p.(Arg429Profs*72);tmVar:p|FS|R,P|429|RO|72;HGVS:p.R,P429ROfsX72;VariantGroup:28;CorrespondingGene:582

p.(Asn354Lys);tmVar:p|SUB|N|354|K;HGVS:p.N354K;VariantGroup:23;CorrespondingGene:583

0no label

Five anencephaly cases carried rare or novel @GENE$ missense variants, three of whom carried additional rare potentially damaging PCP variants: 01F377 (CELSR1 c.6362G>A and @GENE$ c.730C>G), 2F07 (CELSR1 c.8807C>T and DVL3 @VARIANT$), 618F05 (CELSR1 c.8282C>T and SCRIB c.3979G>A). One patient (f93-80) had a novel PTK7 missense variant (c.655A>G) with a rare CELSR2 missense variant (c.1892C>T). Three patients carried missense variants both in FZD and other PCP-associated genes: 01F552 (FZD6 c.1531C>T and CELSR2 c.3800A>G), 335F07 (FZD6 c.544G>A and 2 FAT4 missense variants @VARIANT$; c.10384A>G), and 465F99 (rare FZD1 missense variant c.211C>T and a novel FAT4 missense variant c.10147G>A).

CELSR1;7665

PRICKLE4;22752

c.1622C>T;tmVar:c|SUB|C|1622|T;HGVS:c.1622C>T;VariantGroup:5;CorrespondingGene:1857;RS#:1311053970

c.5792A>G;tmVar:c|SUB|A|5792|G;HGVS:c.5792A>G;VariantGroup:2;CorrespondingGene:79633;RS#:373263457;CA#:4677776

0no label

(D) SH175-389 harbored a monoallelic @VARIANT$ variant of GJB2 and a monoallelic @VARIANT$ variant of GJB3. DFNB1 = nonsyndromic hearing loss and deafness 1, GJB2 = @GENE$, @GENE$ = gap junction protein beta 3, GJB6 = gap junction protein beta 6, MITF = microphthalmia-associated transcription factor.

gap junction protein beta 2;2975

GJB3;7338

p.V193E;tmVar:p|SUB|V|193|E;HGVS:p.V193E;VariantGroup:21;CorrespondingGene:2706

p.A194T;tmVar:p|SUB|A|194|T;HGVS:p.A194T;VariantGroup:18;CorrespondingGene:2707;RS#:117385606;CA#:118313

0no label

Results Cosegregating deleterious variants (GRCH37/hg19) in CACNA1A (NM_001127222.1: c.7261_7262delinsGT, p.Pro2421Val), REEP4 (NM_025232.3: c.109C>T, p.Arg37Trp), TOR2A (NM_130459.3: c.568C>T, @VARIANT$), and ATP2A3 (NM_005173.3: c.1966C>T, @VARIANT$) were identified in four independent multigenerational pedigrees. Deleterious variants in HS1BP3 (NM_022460.3: c.94C>A, p.Gly32Cys) and GNA14 (NM_004297.3: c.989_990del, p.Thr330ArgfsTer67) were identified in a father and son with segmental cranio-cervical dystonia first manifest as BSP. Deleterious variants in DNAH17,@GENE$,@GENE$,VPS13C,UNC13B,SPTBN4,MYOD1, and MRPL15 were found in two or more independent pedigrees.

TRPV4;11003

CAPN11;21392

p.Arg190Cys;tmVar:p|SUB|R|190|C;HGVS:p.R190C;VariantGroup:12;CorrespondingGene:27433;RS#:376074923;CA#:5250615

p.Arg656Cys;tmVar:p|SUB|R|656|C;HGVS:p.R656C;VariantGroup:21;CorrespondingGene:489;RS#:140404080;CA#:8297011

0no label

Co-segregation of TEK p.I148T and CYP1B1 @VARIANT$ was observed in two pedigrees and only a representative pedigree is shown. b Chromatograms of the four probands (lower panel) harboring the four different heterozygous @GENE$ mutations. The site of nucleotide change is indicated by an arrow, compared to the corresponding wild-type sequence (upper panel). c TEK protein sequence conservation across different species for the four mutations (p.E103D, @VARIANT$, p.Q214P, and p.G743A). The conserved residue for each mutation is highlighted in blue color. d Schematic representation of the TEK and CYP1B1 domains (Ig immunoglobulin, EGF epidermal growth factor, FN fibronectin, TM transmembrane, M membrane, H hinge region) indicating the location of the mutations identified in PCG (color figure online) TEK and @GENE$ interact in cells.

TEK;397

CYP1B1;68035

p.R368H;tmVar:p|SUB|R|368|H;HGVS:p.R368H;VariantGroup:1;CorrespondingGene:1545;RS#:79204362;CA#:119016

p.I148T;tmVar:p|SUB|I|148|T;HGVS:p.I148T;VariantGroup:5;CorrespondingGene:7010;RS#:35969327;CA#:5015918

0no label

CONCLUSIONS We firstly identified the novel digenic heterozygous mutations by WES, @GENE$ @VARIANT$ and @GENE$ @VARIANT$, which resulted in LQTS with repeat syncope, torsades de pointes, ventricular fibrillation, and sinoatrial node dysfunction.

KCNH2;201

SCN5A;22738

p.307_308del;tmVar:p|DEL|307_308|;HGVS:p.307_308del;VariantGroup:16;CorrespondingGene:3757

p.R1865H;tmVar:p|SUB|R|1865|H;HGVS:p.R1865H;VariantGroup:1;CorrespondingGene:6331;RS#:370694515;CA#:64651

11

Amino acid conservation analysis showed that seven of the 10 variants (CELSR1 p.G1122S, CELSR1 p.R769W, DVL3 p.R148Q, PTK7 p.P642R, @GENE$ p.G1108E, SCRIB p.G644V and SCRIB p.K618R) were located at highly conserved nucleotides in human, dog, mouse, rat, and zebrafish. The four other variants (CELSR1 @VARIANT$, CELSR1 p.R1057C and SCRIB p.R1044Q) involved less conserved nucleotides (Supplemental material, Fig. S2). Among these variants, p.R769W and p.R1057C localized to the carbonic anhydrases subunits, named the CA domain of @GENE$, p.R1044Q was within the third PDZ domain of SCRIB, @VARIANT$ located very close to the fourth PDZ domain (1109-1192) of SCRIB, and p.P642R was within the fifth IGc2 domain of PTK7 (Supplemental Material, Fig. S3).

SCRIB;44228

CELSR1;7665

p.Q2924H;tmVar:p|SUB|Q|2924|H;HGVS:p.Q2924H;VariantGroup:1;CorrespondingGene:9620;RS#:200116798;CA#:10292663

p.G1108E;tmVar:p|SUB|G|1108|E;HGVS:p.G1108E;VariantGroup:3;CorrespondingGene:23513;RS#:529610993;CA#:4918763

0no label

In patient AVM028, one novel heterozygous VUS (c.2207A>G [@VARIANT$]) in @GENE$ inherited from the father and one likely pathogenic de novo novel heterozygous variant (c.311T>C [@VARIANT$]) in TIMP3 were identified (online supplementary table S2). While @GENE$ blocks VEGF/VEGFR2 signalling, RASA1 modulates differentiation and proliferation of blood vessel endothelial cells downstream of VEGF (figure 3).

RASA1;2168

TIMP3;36322

p.His736Arg;tmVar:p|SUB|H|736|R;HGVS:p.H736R;VariantGroup:6;CorrespondingGene:5921;RS#:1403332745

p.Leu104Pro;tmVar:p|SUB|L|104|P;HGVS:p.L104P;VariantGroup:7;CorrespondingGene:23592;RS#:1290872293

0no label

SCAP-c.3035C>T (p.Ala1012Val) variant impaired the negative feedback mechanism of cholesterol synthesize in H293T cell lines SCAP-c.3035C>T (@VARIANT$) variant was introduced into H293T cell lines by CRISPR-Cas9 methodology. After incubated with medium A (as described in the materials and methods section) for 6 hours, the wild-type goups showed a significant different distribution of SREBP-2 in cytoplasm and nucleus, (Figure 4A) while the @GENE$-mutated groups shows no such difference (Figure 4B). These phenomenon indicate that the mutated SCAP-c.3035C>T (p.Ala1012Val) protein failed to sensing the intracellular cholesterol level, implying a loss of negative feedback mechanism of the mutated SCAP coding protein. AGXT2-c.1103C>T (p.Ala338Val) variant impaired the catabolism of ADMA in EA. hy926 cell lines @GENE$-@VARIANT$ (p.Ala338Val) variant was introduced into EA.

SCAP;8160

AGXT2;12887

p.Ala1012Val;tmVar:p|SUB|A|1012|V;HGVS:p.A1012V;VariantGroup:2;CorrespondingGene:22937

c.1103C>T;tmVar:c|SUB|C|1103|T;HGVS:c.1103C>T;VariantGroup:3;CorrespondingGene:64902;RS#:536786734;CA#:116921745

0no label

We identified a novel compound heterozygous variant in BBS1 @VARIANT$ (p.(Arg429Profs*72); a likely pathogenic novel variant affecting the conserved residue 354 in the functional domain of BBS2 (c.1062C > G; p.(Asn354Lys)); a pathogenic new homozygous nucleotide change in @GENE$ that leads to a stop codon in position 255, c.763A > T, and a likely pathogenic homozygous substitution @VARIANT$ in @GENE$, leading to the change p.(Cys412Phe).

BBS7;12395

BBS6;10318

c.1285dup;tmVar:c|DUP|1285||;HGVS:c.1285dup;VariantGroup:20;CorrespondingGene:582

c.1235G > T;tmVar:c|SUB|G|1235|T;HGVS:c.1235G>T;VariantGroup:15;CorrespondingGene:8195;RS#:1396840386

0no label

The T338I and @VARIANT$ variants affect the conserved central coiled-coil rod domain of the protein mediating dimerization; therefore, we suggest their potential deleterious effect on the protein. In the individual carrying the @VARIANT$ NEFH variant, an additional novel alteration (C335R) was detected in the GRN gene. Loss-of-function GRN variants are primarily considered to cause frontotemporal lobar degeneration, but there is evidence that missense @GENE$ variants are also linked to the pathogenesis of ALS. The novel GRN variant reported in this study results in a cysteine-to-arginine change in the cysteine-rich granulin A domain. Four cases were identified to carry SQSTM1 variants: the P392L in two cases and the E389Q and R393Q in single patients. All three alterations are located within the C-terminal ubiquitin-associated (UBA) end of the sequestome 1 protein. Variants of the @GENE$ gene were originally reported in Paget's disease of bone.

GRN;1577

SQSTM1;31202

R148P;tmVar:p|SUB|R|148|P;HGVS:p.R148P;VariantGroup:14;CorrespondingGene:2521;RS#:773655049

P505L;tmVar:p|SUB|P|505|L;HGVS:p.P505L;VariantGroup:22;CorrespondingGene:4744;RS#:1414968372

0no label

We identified a novel compound heterozygous variant in BBS1 c.1285dup (p.(Arg429Profs*72); a likely pathogenic novel variant affecting the conserved residue 354 in the functional domain of @GENE$ (c.1062C > G; p.(Asn354Lys)); a pathogenic new homozygous nucleotide change in @GENE$ that leads to a stop codon in position 255, @VARIANT$, and a likely pathogenic homozygous substitution c.1235G > T in BBS6, leading to the change p.(@VARIANT$).

BBS2;12122

BBS7;12395

c.763A > T;tmVar:c|SUB|A|763|T;HGVS:c.763A>T;VariantGroup:29;CorrespondingGene:55212

Cys412Phe;tmVar:p|SUB|C|412|F;HGVS:p.C412F;VariantGroup:15;CorrespondingGene:8195;RS#:1396840386

0no label

Based on both clinical and laboratory quantification, it appears neither the @GENE$/TACI @VARIANT$ mutation nor the @GENE$ @VARIANT$ mutation alone is sufficient to cause the complete, severe CVID-like disorder and SLE observed in the proband.

TNFRSF13B;49320

TCF3;2408

C104R;tmVar:p|SUB|C|104|R;HGVS:p.C104R;VariantGroup:2;CorrespondingGene:23495;RS#:34557412;CA#:117387

T168fsX191;tmVar:p|FS|T|168||191;HGVS:p.T168fsX191;VariantGroup:1;CorrespondingGene:6929

11

Two unrelated KS patients had heterozygous NELF mutations and mutation in a second gene: NELF/KAL1 (c.757G>A; p.Ala253Thr of NELF and c.488_490delGTT; @VARIANT$ of @GENE$) and @GENE$/TACR3 (c. 1160-13C>T of NELF and @VARIANT$; p.Trp275X of TACR3).

KAL1;55445

NELF;10648

p.Cys163del;tmVar:p|DEL|163|C;HGVS:p.163delC;VariantGroup:10;CorrespondingGene:3730

c.824G>A;tmVar:c|SUB|G|824|A;HGVS:c.824G>A;VariantGroup:1;CorrespondingGene:26012;RS#:144292455;CA#:144871

0no label

Compared to WT (wild-type) proteins, we found that the ability of GFP-CYP1B1 A115P and GFP-CYP1B1 E229K to immunoprecipitate HA-@GENE$ @VARIANT$ and HA-TEK Q214P, respectively, was significantly diminished. GFP-@GENE$ @VARIANT$ also exhibited relatively reduced ability to immunoprecipitate HA-TEK I148T (~70%).

TEK;397

CYP1B1;68035

E103D;tmVar:p|SUB|E|103|D;HGVS:p.E103D;VariantGroup:2;CorrespondingGene:7010;RS#:572527340;CA#:5015873

R368H;tmVar:p|SUB|R|368|H;HGVS:p.R368H;VariantGroup:1;CorrespondingGene:1545;RS#:79204362;CA#:119016

0no label

SCN5A @VARIANT$ and @GENE$ p.307_308 of amino acid sequences were highly conserved across the common species Sanger sequencing for @GENE$ and KCNH2 mutations. KCNH2 @VARIANT$ and SCN5A p.R1865H of the proband were validated as positive by Sanger sequencing.

KCNH2;201

SCN5A;22738

p.R1865;tmVar:p|Allele|R|1865;VariantGroup:1;CorrespondingGene:6331;RS#:370694515

p.307_308del;tmVar:p|DEL|307_308|;HGVS:p.307_308del;VariantGroup:16;CorrespondingGene:3757

0no label

(C) Sanger sequencing confirmed a homozygous in-frame deletion (@VARIANT$) in MYD88 gene and a homozygous splice-donor mutation (@VARIANT$) in @GENE$ gene. (D) Western Blot of CARD9 and @GENE$ proteins performed on PBMC, EBVB, and PHA derived T cell lines.

CARD9;14150

MYD88;1849

c.195_197delGGA;tmVar:p|DEL|195_197|G;HGVS:p.195_197delG;VariantGroup:2;CorrespondingGene:4615

c.1434+1G>C;tmVar:c|SUB|G|1434+1|C;HGVS:c.1434+1G>C;VariantGroup:0;CorrespondingGene:64170;RS#:141992399;CA#:500026

0no label

Therefore, in this study, SCN5A p.R1865H may be the main cause of sinoatrial node dysfunction, whereas KCNH2 @VARIANT$ only carried by II: 1 may potentially induce the phenotype of LQTS. However, it was hard to determine whether the coexisting interactions of @GENE$ p.307_308del and SCN5A p.R1865H increased the risk of young and early-onset LQTS, or whether KCNH2 mutation was only associated with LQTS, while SCN5A mutation was only associated with sinoatrial node dysfunction. CONCLUSIONS We firstly identified the novel digenic heterozygous mutations by WES, KCNH2 p.307_308del and @GENE$ p.R1865H, which resulted in LQTS with repeat syncope, torsades de pointes, ventricular fibrillation, and sinoatrial node dysfunction. KCNH2 p.307_308del may affect the function of Kv11.1 channel in cardiomyocytes by inducing a regional double helix of the amino acids misfolded and largest hydrophobic domain disorganized. SCN5A @VARIANT$ reduced the instability index of Nav1.5 protein and sodium current.

KCNH2;201

SCN5A;22738

p.307_308del;tmVar:p|DEL|307_308|;HGVS:p.307_308del;VariantGroup:16;CorrespondingGene:3757

p.R1865H;tmVar:p|SUB|R|1865|H;HGVS:p.R1865H;VariantGroup:1;CorrespondingGene:6331;RS#:370694515;CA#:64651

0no label

(A) In the @GENE$ exon 4 and exon 9, the arrows indicate the nucleotide substitution, c.475A > G and @VARIANT$, consisting, respectively, in the amino acid substitutions, S159G (A/G heterozygous patient and mother, A/A wild type father) and R351G; (B) in the @GENE$ exon 9 sequence, the c.2857 A > G substitution consisted in an amino acid substitution, K953E (A/G heterozygous patient and mother, A/A wild-type father). Bioinformatics analysis results. (A) Multiple alignment of the amino acid sequence of NOD2 protein in seven species showed that this is a conserved region; (B) PolyPhen2 (Polymorphism Phenotyping v.2) analysis predicting the probably damaging impact of the @VARIANT$ substitution with a score of 0.999.

IL10RA;1196

NOD2;11156

c.1051A > G;tmVar:c|SUB|A|1051|G;HGVS:c.1051A>G;VariantGroup:0;CorrespondingGene:3587;RS#:8178561;CA#:10006322

K953E;tmVar:p|SUB|K|953|E;HGVS:p.K953E;VariantGroup:0;CorrespondingGene:64127;RS#:8178561

0no label

The nucleotide sequence showed a @VARIANT$ (c.769G>C) of the coding sequence in exon 7 of EDA, which results in the substitution of Gly at residue 257 to Arg. Additionally, the nucleotide sequence showed a monoallelic C to T transition at nucleotide 511 (c.511C>T) of the coding sequence in exon 3 of WNT10A, which results in the substitution of @VARIANT$. DNA sequencing of the parents' genome revealed that both mutant alleles were from their mother (Fig. 2A), who carried a heterozygous @GENE$ mutation (c.769G>C) and a heterozygous @GENE$ c.511C>T mutation, and showed absence of only the left upper lateral incisor without other clinical abnormalities.

EDA;1896

WNT10A;22525

G to C transition at nucleotide 769;tmVar:c|SUB|G|769|C;HGVS:c.769G>C;VariantGroup:0;CorrespondingGene:1896;RS#:1057517882;CA#:16043329

Arg at residue 171 to Cys;tmVar:p|SUB|R|171|C;HGVS:p.R171C;VariantGroup:3;CorrespondingGene:80326;RS#:116998555;CA#:2113955

0no label

The nucleotide sequence showed a @VARIANT$ (c.769G>C) of the coding sequence in exon 7 of EDA, which results in the substitution of Gly at residue 257 to Arg. Additionally, the nucleotide sequence showed a monoallelic C to T transition at nucleotide 511 (c.511C>T) of the coding sequence in exon 3 of WNT10A, which results in the substitution of Arg at residue 171 to Cys. DNA sequencing of the parents' genome revealed that both mutant alleles were from their mother (Fig. 2A), who carried a heterozygous EDA mutation (c.769G>C) and a heterozygous WNT10A @VARIANT$ mutation, and showed absence of only the left upper lateral incisor without other clinical abnormalities. No mutations in these genes were found in the father. Sequence analyses of EDA and @GENE$ genes. (A) The @GENE$ mutation c.769G>C and WNT10A mutation c.511C>T were found in patient N1, who inherited the mutant allele from his mother.

WNT10A;22525

EDA;1896

G to C transition at nucleotide 769;tmVar:c|SUB|G|769|C;HGVS:c.769G>C;VariantGroup:0;CorrespondingGene:1896;RS#:1057517882;CA#:16043329

c.511C>T;tmVar:c|SUB|C|511|T;HGVS:c.511C>T;VariantGroup:3;CorrespondingGene:80326;RS#:116998555;CA#:2113955

0no label

We identified a novel variant in the @GENE$ gene (c.2857A > G @VARIANT$) and two already described missense variants in the @GENE$ gene (S159G and @VARIANT$).

NOD2;11156

IL10RA;1196

p.K953E;tmVar:p|SUB|K|953|E;HGVS:p.K953E;VariantGroup:0;CorrespondingGene:64127;RS#:8178561

G351R;tmVar:p|SUB|G|351|R;HGVS:p.G351R;VariantGroup:0;CorrespondingGene:3587;RS#:8178561

0no label

@GENE$ and DSCAM have similar neurodevelopmental functions and are essential for self-avoidance in the developing mouse retina. In patient AVM144, the compound heterozygous variants @VARIANT$ and @VARIANT$ (p.Ser334Thr) were identified in @GENE$ (table 2).

DSCAML1;79549

PTPN13;7909

c.116-1G>A;tmVar:c|SUB|G|116-1|A;HGVS:c.116-1G>A;VariantGroup:5;CorrespondingGene:83394;RS#:1212415588

c.1000T>A;tmVar:c|SUB|T|1000|A;HGVS:c.1000T>A;VariantGroup:0;CorrespondingGene:5783;RS#:755467869;CA#:2995566

0no label

The @VARIANT$ (c.1045G>A) mutation in exon 9 of @GENE$ and heterozygous p.Arg171Cys (c.511C>T) mutation in exon 3 of @GENE$ were detected. These mutations were not found in his father's genome, but because his mother's DNA sample was unavailable, the origin of the mutant alleles was not clear (Fig. 2F). All novel mutations that were identified in this study were not found in the normal controls. Protein structure analysis The results of protein structure analyses of WNT10A are shown in Figure 3. R171 and G213 are conserved residues through these organisms and located on conserved 2D fragments. Mutations of the residues could affect the function of the human WNT10A protein. In the case of R171C mutations, the substitution of Cys, a hydroxylic amino acid with a side chain shorter than Arg, might eliminate the electrostatic interaction of R171 with adjacent residues. The mutation G213S is expected to abolish the hydrophobic interaction of @VARIANT$ with adjacent residues.

EDA;1896

WNT10A;22525

p.Ala349Thr;tmVar:p|SUB|A|349|T;HGVS:p.A349T;VariantGroup:2;CorrespondingGene:1896;RS#:132630317;CA#:255657

G213;tmVar:c|Allele|G|213;VariantGroup:4;CorrespondingGene:80326;RS#:147680216

0no label

WES revealed heterozygous mutations in two genes known to affect hypothalamic and pituitary development: @VARIANT$;p.R85C in @GENE$ (MIM 607123; NM_144773.2; rs141090506) inherited from an unaffected mother and c.1306A>G;@VARIANT$ in @GENE$ (MIM 606417; NM_018117.11; rs34602786) inherited from an unaffected father, both confirmed by Sanger sequencing (Fig. 1).

PROKR2;16368

WDR11;41229

c.253C>T;tmVar:c|SUB|C|253|T;HGVS:c.253C>T;VariantGroup:1;CorrespondingGene:128674;RS#:74315418;CA#:259601

p.I436V;tmVar:p|SUB|I|436|V;HGVS:p.I436V;VariantGroup:3;CorrespondingGene:55717;RS#:34602786;CA#:5719694

11

Hence, priority should be given to identifying the TCF3 @VARIANT$ mutation for preimplantation genetic diagnosis and/or chorionic villus sampling. Based on both clinical and laboratory quantification, it appears neither the @GENE$/TACI @VARIANT$ mutation nor the @GENE$ T168fsX191 mutation alone is sufficient to cause the complete, severe CVID-like disorder and SLE observed in the proband.

TNFRSF13B;49320

TCF3;2408

T168fsX191;tmVar:p|FS|T|168||191;HGVS:p.T168fsX191;VariantGroup:1;CorrespondingGene:6929

C104R;tmVar:p|SUB|C|104|R;HGVS:p.C104R;VariantGroup:2;CorrespondingGene:23495;RS#:34557412;CA#:117387

0no label

Exome analysis for the proband identified three sequence variants in FTA candidate genes, two in @GENE$ (g.27546T>A, c.379T>A, p.Ser127Thr; g.124339A>G, c.3224A>G, p.Asn1075Ser) and one in @GENE$ (g.14574G>C, @VARIANT$, p.Glu167Gln) (Figure 4A). The LRP6 @VARIANT$ mutation is a rare variant with an MAF of 0.0024 in EAS.

LRP6;1747

WNT10A;22525

c.499G>C;tmVar:c|SUB|G|499|C;HGVS:c.499G>C;VariantGroup:5;CorrespondingGene:80326;RS#:148714379

c.3224A>G;tmVar:c|SUB|A|3224|G;HGVS:c.3224A>G;VariantGroup:8;CorrespondingGene:4040;RS#:202124188

0no label

The nucleotide sequence showed a G to C transition at nucleotide 769 (@VARIANT$) of the coding sequence in exon 7 of EDA, which results in the substitution of Gly at residue 257 to Arg. Additionally, the nucleotide sequence showed a monoallelic C to T transition at nucleotide 511 (c.511C>T) of the coding sequence in exon 3 of @GENE$, which results in the substitution of Arg at residue 171 to Cys. DNA sequencing of the parents' genome revealed that both mutant alleles were from their mother (Fig. 2A), who carried a heterozygous EDA mutation (c.769G>C) and a heterozygous WNT10A @VARIANT$ mutation, and showed absence of only the left upper lateral incisor without other clinical abnormalities. No mutations in these genes were found in the father. Sequence analyses of EDA and WNT10A genes. (A) The @GENE$ mutation c.769G>C and WNT10A mutation c.511C>T were found in patient N1, who inherited the mutant allele from his mother.

WNT10A;22525

EDA;1896

c.769G>C;tmVar:c|SUB|G|769|C;HGVS:c.769G>C;VariantGroup:0;CorrespondingGene:1896;RS#:1057517882;CA#:16043329

c.511C>T;tmVar:c|SUB|C|511|T;HGVS:c.511C>T;VariantGroup:3;CorrespondingGene:80326;RS#:116998555;CA#:2113955

0no label

Because charged residues are important for proteins trafficking, the @VARIANT$ may result in accumulation of the Cx31 protein in intracellular compartments such as the Golgi apparatus or in other sites such as the endoplasmic reticulum or lysosomes. The A194T substitution might cause conformational changes within the Cx31 molecule or affect the ability of @GENE$ to form heteromeric hemichannels. The relationship between hemichannel assembly may be complex, considering the different paradigms for connexin oligomerization. Many of the Cx26 mutant residues lie within the EC2 and TM4 domains. Mutations affecting these regions have also been reported in Cx32 underlying X-linked-Charcot-Marie-Tooth disease. Moreover, mutations in residues close to N166 and A194 identified in the families reported here, namely, M163L, R165W, F191L, and A197S in Cx26 as well as @VARIANT$, S198F and G199R in @GENE$, have been reported previously in patients with hearing impairment.

Cx31;7338

Cx32;137

A194T;tmVar:c|SUB|A|194|T;HGVS:c.194A>T;VariantGroup:4;CorrespondingGene:2707;RS#:117385606;CA#:118313

F193C;tmVar:p|SUB|F|193|C;HGVS:p.F193C;VariantGroup:15;CorrespondingGene:2706

0no label

The @VARIANT$ nonsense variant was first detected in compound heterozygous form in a family with two affected siblings suffering from infantile ascending spastic paralysis with bulbar involvement. The ages of onset of the patients with the ALS2 variants reported in this study were later than juvenile ALS onset, which generally manifests before 25 years of age. Previous studies suggested that heterozygous variants in the @GENE$ may be causative for adult-onset sALS. MATR3 encodes three protein isoforms that have been described as nuclear-matrix and DNA/RNA binding proteins involved in transcription and stabilization of mRNA. In the present study, two novel heterozygous variants (P11S, @VARIANT$) were detected. The P11S variant affects the b isoform of the @GENE$ protein (NM_001194956 and NP_001181885), contributing to splicing alteration of other isoforms.

ALS2;23264

MATR3;7830

G1177X;tmVar:p|SUB|G|1177|X;HGVS:p.G1177X;VariantGroup:0;CorrespondingGene:57679;RS#:386134180;CA#:356568

S275N;tmVar:p|SUB|S|275|N;HGVS:p.S275N;VariantGroup:9;CorrespondingGene:80208;RS#:995711809

0no label

21 Additional gene reportedly linked to tumorigenesis include RYR3, 22 @GENE$, 23 @GENE$, 24 and CAPN9. 25 The RYR3 (NM_001036: @VARIANT$, p.Asn2604Lys) and EBNA1BP2 (NM_001159936: c.1034A > T, p.Asn345Ile) variants were classified as likely benign and benign, respectively, while the TRIP6 (NM_003302: c.822G > C, p.Glu274Asp) and the CAPN9 (NM_006615: c.55G > T, @VARIANT$) variants were classified as VUS.

EBNA1BP2;4969

TRIP6;37757

c.7812C > G;tmVar:c|SUB|C|7812|G;HGVS:c.7812C>G;VariantGroup:10;CorrespondingGene:6263;RS#:41279214;CA#:7459988

p.Ala19Ser;tmVar:p|SUB|A|19|S;HGVS:p.A19S;VariantGroup:17;CorrespondingGene:10753;RS#:147360179;CA#:1448452

0no label

This analysis indicated that the @GENE$ variant @VARIANT$ (rs138172448), which results in a p.Val555Ile change, and the @GENE$ gene variant c.656C>T (@VARIANT$), which results in a p.Thr219Ile change, are both predicted to be damaging.

CAPN3;52

DES;56469

c.1663G>A;tmVar:c|SUB|G|1663|A;HGVS:c.1663G>A;VariantGroup:2;CorrespondingGene:825;RS#:138172448;CA#:7511461

rs144901249;tmVar:rs144901249;VariantGroup:3;CorrespondingGene:1674;RS#:144901249

0no label

In our study, we identified four genetic variants in three genes (@GENE$-p.R583H, @GENE$-@VARIANT$, KCNH2-p.K897T, and KCNE1-@VARIANT$).

KCNQ1;85014

KCNH2;201

p.C108Y;tmVar:p|SUB|C|108|Y;HGVS:p.C108Y;VariantGroup:3;CorrespondingGene:3757

p.G38S;tmVar:p|SUB|G|38|S;HGVS:p.G38S;VariantGroup:1;CorrespondingGene:3753;RS#:1805127;CA#:131330

0no label

In our study, we identified four genetic variants in three genes (@GENE$-p.R583H, KCNH2-p.C108Y, @GENE$-@VARIANT$, and KCNE1-@VARIANT$).

KCNQ1;85014

KCNH2;201

p.K897T;tmVar:p|SUB|K|897|T;HGVS:p.K897T;VariantGroup:0;CorrespondingGene:3757;RS#:1805123;CA#:7162

p.G38S;tmVar:p|SUB|G|38|S;HGVS:p.G38S;VariantGroup:1;CorrespondingGene:3753;RS#:1805127;CA#:131330

0no label

Two different @GENE$ mutations (N166S and @VARIANT$) occurring in compound heterozygosity with the 235delC and 299delAT of @GENE$ were identified in three unrelated families (235delC/N166S, @VARIANT$/A194T and 299delAT/A194T).

GJB3;7338

GJB2;2975

A194T;tmVar:c|SUB|A|194|T;HGVS:c.194A>T;VariantGroup:4;CorrespondingGene:2707;RS#:117385606;CA#:118313

235delC;tmVar:c|DEL|235|C;HGVS:c.235delC;VariantGroup:1;CorrespondingGene:2706;RS#:80338943

0no label

Sanger sequencing of Family 1 showed that both rs138355706 in @GENE$ (@VARIANT$, missense causing a p.R77C mutation) and a 4 bp deletion in @GENE$ (c.238-241delATTG causing a frameshift @VARIANT$) segregated completely with ILD in Family 1 based upon recessive inheritance (figure 2c and d), were in total linkage disequilibrium, and were present in a cis conformation.

S100A3;2223

S100A13;7523

c.229C>T;tmVar:c|SUB|C|229|T;HGVS:c.229C>T;VariantGroup:3;CorrespondingGene:6274;RS#:138355706;CA#:1116284

p.I80Gfs*13;tmVar:p|FS|I|80|G|13;HGVS:p.I80GfsX13;VariantGroup:7;CorrespondingGene:6284

11

Missense variants in the NEFH gene were detected in four patients: the T338I variant in two cases and the R148P and @VARIANT$ variants in single cases. NEFH encodes the heavy neurofilament protein, and its variants have been associated with neuronal damage in ALS. The T338I and R148P variants affect the conserved central coiled-coil rod domain of the protein mediating dimerization; therefore, we suggest their potential deleterious effect on the protein. In the individual carrying the P505L NEFH variant, an additional novel alteration (@VARIANT$) was detected in the GRN gene. Loss-of-function GRN variants are primarily considered to cause frontotemporal lobar degeneration, but there is evidence that missense GRN variants are also linked to the pathogenesis of ALS. The novel @GENE$ variant reported in this study results in a cysteine-to-arginine change in the cysteine-rich granulin A domain. Four cases were identified to carry @GENE$ variants: the P392L in two cases and the E389Q and R393Q in single patients.

GRN;1577

SQSTM1;31202

P505L;tmVar:p|SUB|P|505|L;HGVS:p.P505L;VariantGroup:22;CorrespondingGene:4744;RS#:1414968372

C335R;tmVar:p|SUB|C|335|R;HGVS:p.C335R;VariantGroup:29;CorrespondingGene:29110;RS#:1383907519

0no label

Two different GJB3 mutations (N166S and A194T) occurring in compound heterozygosity with the @VARIANT$ and 299delAT of @GENE$ were identified in three unrelated families (235delC/@VARIANT$, 235delC/A194T and 299delAT/A194T). Neither of these mutations in @GENE$ was detected in DNA from 200 unrelated Chinese controls.

GJB2;2975

Cx31;7338

235delC;tmVar:c|DEL|235|C;HGVS:c.235delC;VariantGroup:1;CorrespondingGene:2706;RS#:80338943

N166S;tmVar:p|SUB|N|166|S;HGVS:p.N166S;VariantGroup:0;CorrespondingGene:2707;RS#:121908851;CA#:118311

0no label

Recently, rare heterozygous alleles in the angiopoietin receptor-encoding gene @GENE$ were implicated in PCG. We undertook this study to ascertain the second mutant allele in a large cohort (n = 337) of autosomal recessive PCG cases that carried heterozygous TEK mutations. Our investigations revealed 12 rare heterozygous missense mutations in TEK by targeted sequencing. Interestingly, four of these TEK mutations (p.E103D, @VARIANT$, p.Q214P, and p.G743A) co-occurred with three heterozygous mutations in another major PCG gene @GENE$ (@VARIANT$, p.E229K, and p.R368H) in five families.

TEK;397

CYP1B1;68035

p.I148T;tmVar:p|SUB|I|148|T;HGVS:p.I148T;VariantGroup:5;CorrespondingGene:7010;RS#:35969327;CA#:5015918

p.A115P;tmVar:p|SUB|A|115|P;HGVS:p.A115P;VariantGroup:0;CorrespondingGene:1545;RS#:764338357;CA#:1620052

0no label

Two unrelated KS patients had heterozygous NELF mutations and mutation in a second gene: NELF/@GENE$ (c.757G>A; @VARIANT$ of NELF and c.488_490delGTT; p.Cys163del of KAL1) and NELF/TACR3 (c. 1160-13C>T of @GENE$ and c.824G>A; @VARIANT$ of TACR3).

KAL1;55445

NELF;10648

p.Ala253Thr;tmVar:p|SUB|A|253|T;HGVS:p.A253T;VariantGroup:3;CorrespondingGene:26012;RS#:142726563;CA#:5370407

p.Trp275X;tmVar:p|SUB|W|275|X;HGVS:p.W275X;VariantGroup:1;CorrespondingGene:6870;RS#:144292455;CA#:144871

0no label

Subsequently many genes encoding folate pathway enzymes, transporters and receptors have been studied with mostly inconsistent findings.7 More recently, several candidate variants were identified in @GENE$ and @GENE$, 2 of the genes constituting the mitochondrial GCS.10, 32 In the present study, we identified a novel missense variant affecting the catalytic domain of the MTHFR gene. This patient additionally carried the @VARIANT$ variant, and a rare missense variant (@VARIANT$) in the GLDC gene.

AMT;409

GLDC;141

c.677C>T;tmVar:c|SUB|C|677|T;HGVS:c.677C>T;VariantGroup:27;CorrespondingGene:4524;RS#:1801133;CA#:170990

c.2203G>T;tmVar:c|SUB|G|2203|T;HGVS:c.2203G>T;VariantGroup:3;CorrespondingGene:2731;RS#:143119940;CA#:4980332

0no label

The ISG20L2 and @GENE$ variants were excluded based on their frequencies in normal population cohorts. Sanger sequencing of Family 1 showed that both rs138355706 in S100A3 (@VARIANT$, missense causing a p.R77C mutation) and a 4 bp deletion in @GENE$ (@VARIANT$ causing a frameshift p.I80Gfs*13) segregated completely with ILD in Family 1 based upon recessive inheritance (figure 2c and d), were in total linkage disequilibrium, and were present in a cis conformation.

SETDB1;32157

S100A13;7523

c.229C>T;tmVar:c|SUB|C|229|T;HGVS:c.229C>T;VariantGroup:3;CorrespondingGene:6274;RS#:138355706;CA#:1116284

c.238-241delATTG;tmVar:c|DEL|238_241|ATTG;HGVS:c.238_241delATTG;VariantGroup:13;CorrespondingGene:6284

0no label

Amino acid conservation analysis showed that seven of the 10 variants (CELSR1 p.G1122S, CELSR1 p.R769W, DVL3 p.R148Q, @GENE$ p.P642R, SCRIB p.G1108E, SCRIB @VARIANT$ and SCRIB @VARIANT$) were located at highly conserved nucleotides in human, dog, mouse, rat, and zebrafish. The four other variants (CELSR1 p.Q2924H, @GENE$ p.R1057C and SCRIB p.R1044Q) involved less conserved nucleotides (Supplemental material, Fig. S2).

PTK7;43672

CELSR1;7665

p.G644V;tmVar:p|SUB|G|644|V;HGVS:p.G644V;VariantGroup:9;CorrespondingGene:23513;RS#:201104891;CA#:187609256

p.K618R;tmVar:p|SUB|K|618|R;HGVS:p.K618R;VariantGroup:2;CorrespondingGene:5754;RS#:139041676

0no label

Using SIFT and PolyPhen, the c.1777C > G variant in SLC9A6 was predicted to be damaging, but a different variant at the same amino acid, c.1777C > T (@VARIANT$), was found in the ExAC database at a rate of 8.24 x 10-6. A male (ID041), unrelated to ID104, carried heterozygous missense variants c.1513G > A (p.Gly505Ser) in @GENE$ and c.353A > G (@VARIANT$) in @GENE$. He was seen at 7 years and 10 months and, at that time, was severely developmentally delayed in multiple domains (motor, cognitive, and language).

EHMT1;11698

MFSD8;115814

p.Leu593Phe;tmVar:p|SUB|L|593|F;HGVS:p.L593F;VariantGroup:7;CorrespondingGene:10479;RS#:149360465;CA#:10524857

p.Asn118Ser;tmVar:p|SUB|N|118|S;HGVS:p.N118S;VariantGroup:5;CorrespondingGene:256471;RS#:774112195;CA#:3077496

0no label

Variants in all known WS candidate genes (@GENE$, EDNRB, @GENE$, PAX3, SOX10, SNAI2, and TYRO3) were searched and a novel rare heterozygous deletion mutation (@VARIANT$; p.Asn322fs) was identified in the MITF gene in both patients. Moreover, heterozygous missense variants in SNAI3 (c.607C>T; @VARIANT$) and TYRO3 (c.1037T>A; p.Ile346Asn) gene was identified in the exome data of both patients.

EDN3;88

MITF;4892

c.965delA;tmVar:c|DEL|965|A;HGVS:c.965delA;VariantGroup:4;CorrespondingGene:4286

p.Arg203Cys;tmVar:p|SUB|R|203|C;HGVS:p.R203C;VariantGroup:1;CorrespondingGene:333929;RS#:149676512;CA#:8229366

0no label

@GENE$ = nonsyndromic hearing loss and deafness 1, GJB2 = gap junction protein beta 2, @GENE$ = gap junction protein beta 3, GJB6 = gap junction protein beta 6, MITF = microphthalmia-associated transcription factor. By screening other gap junction genes, another subject (SH175-389) carrying a single heterozygous @VARIANT$ in GJB2 allele harbored a single heterozygous @VARIANT$ mutant allele of GJB3 (NM_001005752) (SH175-389) with known pathogenicity (Figure 4D).

DFNB1;2975

GJB3;7338

p.V193E;tmVar:p|SUB|V|193|E;HGVS:p.V193E;VariantGroup:21;CorrespondingGene:2706

p.A194T;tmVar:p|SUB|A|194|T;HGVS:p.A194T;VariantGroup:18;CorrespondingGene:2707;RS#:117385606;CA#:118313

0no label

The nucleotide sequence showed a @VARIANT$ (c.769G>C) of the coding sequence in exon 7 of EDA, which results in the substitution of Gly at residue 257 to Arg. Additionally, the nucleotide sequence showed a monoallelic C to T transition at nucleotide 511 (c.511C>T) of the coding sequence in exon 3 of WNT10A, which results in the substitution of Arg at residue 171 to Cys. DNA sequencing of the parents' genome revealed that both mutant alleles were from their mother (Fig. 2A), who carried a heterozygous EDA mutation (c.769G>C) and a heterozygous WNT10A c.511C>T mutation, and showed absence of only the left upper lateral incisor without other clinical abnormalities. No mutations in these genes were found in the father. Sequence analyses of @GENE$ and WNT10A genes. (A) The EDA mutation c.769G>C and @GENE$ mutation @VARIANT$ were found in patient N1, who inherited the mutant allele from his mother.

EDA;1896

WNT10A;22525

G to C transition at nucleotide 769;tmVar:c|SUB|G|769|C;HGVS:c.769G>C;VariantGroup:0;CorrespondingGene:1896;RS#:1057517882;CA#:16043329

c.511C>T;tmVar:c|SUB|C|511|T;HGVS:c.511C>T;VariantGroup:3;CorrespondingGene:80326;RS#:116998555;CA#:2113955

0no label

We identified a novel variant in the @GENE$ gene (c.@VARIANT$ p.K953E) and two already described missense variants in the @GENE$ gene (@VARIANT$ and G351R).

NOD2;11156

IL10RA;1196

2857A > G;tmVar:c|SUB|A|2857|G;HGVS:c.2857A>G;VariantGroup:0;CorrespondingGene:64127;RS#:8178561;CA#:10006322

S159G;tmVar:p|SUB|S|159|G;HGVS:p.S159G;VariantGroup:0;CorrespondingGene:3587;RS#:8178561

11

RESULTS Mutations at the gap junction proteins @GENE$ and @GENE$ can interact to cause non-syndromic deafness In total, 108 probands screened for mutations in the Cx26 gene were found to carry a single recessive mutant allele. In those samples, no mutation was detected on the second allele either in Cx26-exon-1/splice sites or in GJB6. To investigate the role of GJB3 variations along with GJB2 mutations for a possible combinatory allelic disease inheritance, we have screened patients with heterozygous GJB2 mutations for variants in Cx31 by sequencing. Analysis of the entire coding region of the Cx31 gene revealed the presence of two different missense mutations (N166S and A194T) occurring in compound heterozygosity along with the @VARIANT$ and 299delAT of GJB2 in 3 simplex families (235delC/N166S, 235delC/A194T and 299delAT/A194T). In family A, a profoundly hearing impaired proband was found to be heterozygous for a novel @VARIANT$ of GJB3, resulting in an asparagine into serine substitution in codon 166 (N166S) and for the 235delC of GJB2 (Fig. 1b, d).

Cx26;2975

Cx31;7338

235delC;tmVar:c|DEL|235|C;HGVS:c.235delC;VariantGroup:1;CorrespondingGene:2706;RS#:80338943

A to G transition at nucleotide position 497;tmVar:c|SUB|A|497|G;HGVS:c.497A>G;VariantGroup:0;CorrespondingGene:2707;RS#:121908851;CA#:118311

0no label

The nucleotide sequence showed a G to C transition at nucleotide 769 (c.769G>C) of the coding sequence in exon 7 of EDA, which results in the substitution of @VARIANT$. Additionally, the nucleotide sequence showed a monoallelic C to T transition at nucleotide 511 (c.511C>T) of the coding sequence in exon 3 of WNT10A, which results in the substitution of Arg at residue 171 to Cys. DNA sequencing of the parents' genome revealed that both mutant alleles were from their mother (Fig. 2A), who carried a heterozygous EDA mutation (c.769G>C) and a heterozygous WNT10A c.511C>T mutation, and showed absence of only the left upper lateral incisor without other clinical abnormalities. No mutations in these genes were found in the father. Sequence analyses of @GENE$ and @GENE$ genes. (A) The EDA mutation c.769G>C and WNT10A mutation @VARIANT$ were found in patient N1, who inherited the mutant allele from his mother.

EDA;1896

WNT10A;22525

Gly at residue 257 to Arg;tmVar:p|SUB|G|257|R;HGVS:p.G257R;VariantGroup:0;CorrespondingGene:1896;RS#:1057517882;CA#:16043329

c.511C>T;tmVar:c|SUB|C|511|T;HGVS:c.511C>T;VariantGroup:3;CorrespondingGene:80326;RS#:116998555;CA#:2113955

0no label

There is a splicing site mutation c.1339 + 3A>T in @GENE$, inherited from her mother and a missense mutation c.4421C > T (p. (Thr1474Met)) inherited from her father (Figure 1a). In AS patient IID29, in addition to a glycine substitution (p. (@VARIANT$)) in COL4A3 in the heterozygous state, there was another heterozygous nonsense mutation @VARIANT$ in @GENE$ genes.

COL4A5;133559

COL4A4;20071

Gly1119Asp;tmVar:p|SUB|G|1119|D;HGVS:p.G1119D;VariantGroup:21;CorrespondingGene:1285;RS#:764480728;CA#:2147204

c.5026C > T;tmVar:c|SUB|C|5026|T;HGVS:c.5026C>T;VariantGroup:20;CorrespondingGene:1286

0no label

The patient carried a heterozygous variant of unknown significance in @GENE$, p.(@VARIANT$), defined as likely pathogenic in ClinVar, and a missense variant p.(@VARIANT$) in PMM2, classified as likely pathogenic. Recessive mutations in PMM2 were reported as associated to hyperinsulinemic hypoglycemia (HI) and @GENE$ (Cabezas et al.,).

PKHD1;16336

PKD;55680

His3124Tyr;tmVar:p|SUB|H|3124|Y;HGVS:p.H3124Y;VariantGroup:17;CorrespondingGene:5314

Gly42Arg;tmVar:p|SUB|G|42|R;HGVS:p.G42R;VariantGroup:5;CorrespondingGene:5373;RS#:755402538;CA#:7893895

0no label

Variants in all known WS candidate genes (EDN3, EDNRB, MITF, PAX3, SOX10, SNAI2, and @GENE$) were searched and a novel rare heterozygous deletion mutation (c.965delA; @VARIANT$) was identified in the @GENE$ gene in both patients. Moreover, heterozygous missense variants in SNAI3 (c.607C>T; p.Arg203Cys) and TYRO3 (@VARIANT$; p.Ile346Asn) gene was identified in the exome data of both patients.

TYRO3;4585

MITF;4892

p.Asn322fs;tmVar:p|FS|N|322||;HGVS:p.N322fsX;VariantGroup:3;CorrespondingGene:4286

c.1037T>A;tmVar:c|SUB|T|1037|A;HGVS:c.1037T>A;VariantGroup:2;CorrespondingGene:7301;RS#:12148316;CA#:7494886

0no label

Analysis of the entire coding region of the Cx31 gene revealed the presence of two different missense mutations (@VARIANT$ and A194T) occurring in compound heterozygosity along with the 235delC and 299delAT of @GENE$ in 3 simplex families (235delC/N166S, 235delC/A194T and 299delAT/A194T). In family A, a profoundly hearing impaired proband was found to be heterozygous for a novel A to G transition at nucleotide position 497 of @GENE$, resulting in an asparagine into serine substitution in codon 166 (N166S) and for the @VARIANT$ of GJB2 (Fig. 1b, d).

GJB2;2975

GJB3;7338

N166S;tmVar:p|SUB|N|166|S;HGVS:p.N166S;VariantGroup:0;CorrespondingGene:2707;RS#:121908851;CA#:118311

235delC;tmVar:c|DEL|235|C;HGVS:c.235delC;VariantGroup:1;CorrespondingGene:2706;RS#:80338943

0no label

Deleterious variants in HS1BP3 (NM_022460.3: c.94C>A, @VARIANT$) and @GENE$ (NM_004297.3: @VARIANT$, p.Thr330ArgfsTer67) were identified in a father and son with segmental cranio-cervical dystonia first manifest as BSP. Deleterious variants in DNAH17,@GENE$,CAPN11,VPS13C,UNC13B,SPTBN4,MYOD1, and MRPL15 were found in two or more independent pedigrees.

GNA14;68386

TRPV4;11003

p.Gly32Cys;tmVar:p|SUB|G|32|C;HGVS:p.G32C;VariantGroup:25;CorrespondingGene:64342

c.989_990del;tmVar:c|DEL|989_990|;HGVS:c.989_990del;VariantGroup:16;CorrespondingGene:9630;RS#:750424668;CA#:5094137

0no label

In patient AVM144, the compound heterozygous variants @VARIANT$ and c.1000T>A (@VARIANT$) were identified in @GENE$ (table 2). Potential oligogenic inheritance Variants in more than one gene (at least one likely pathogenic variant) with differing inheritance origin were identified in three patients (figure 1). In patient AVM558, a pathogenic heterozygous variant c.920dupA (p.Asn307LysfsTer27) inherited from the mother was identified in @GENE$. Another de novo novel heterozygous missense variant, c.1694G>A (p.Arg565Gln), was identified in MAP4K4 (online supplementary table S2), which encodes the kinase responsible for phosphorylation of residue T312 in SMAD1 to block its activity in BMP/TGF-beta signalling.

PTPN13;7909

ENG;92

c.116-1G>A;tmVar:c|SUB|G|116-1|A;HGVS:c.116-1G>A;VariantGroup:5;CorrespondingGene:83394;RS#:1212415588

p.Ser334Thr;tmVar:p|SUB|S|334|T;HGVS:p.S334T;VariantGroup:0;CorrespondingGene:5783;RS#:755467869;CA#:2995566

0no label

Five anencephaly cases carried rare or novel CELSR1 missense variants, three of whom carried additional rare potentially damaging PCP variants: 01F377 (CELSR1 @VARIANT$ and PRICKLE4 c.730C>G), 2F07 (CELSR1 c.8807C>T and DVL3 c.1622C>T), 618F05 (@GENE$ c.8282C>T and SCRIB c.3979G>A). One patient (f93-80) had a novel PTK7 missense variant (@VARIANT$) with a rare CELSR2 missense variant (c.1892C>T). Three patients carried missense variants both in FZD and other PCP-associated genes: 01F552 (FZD6 c.1531C>T and CELSR2 c.3800A>G), 335F07 (@GENE$ c.544G>A and 2 FAT4 missense variants c.5792A>G; c.10384A>G), and 465F99 (rare FZD1 missense variant c.211C>T and a novel FAT4 missense variant c.10147G>A).

CELSR1;7665

FZD6;2617

c.6362G>A;tmVar:c|SUB|G|6362|A;HGVS:c.6362G>A;VariantGroup:33;CorrespondingGene:9620;RS#:765148329;CA#:10293808

c.655A>G;tmVar:c|SUB|A|655|G;HGVS:c.655A>G;VariantGroup:2;CorrespondingGene:5754;RS#:373263457;CA#:4677776

0no label

Five anencephaly cases carried rare or novel CELSR1 missense variants, three of whom carried additional rare potentially damaging PCP variants: 01F377 (CELSR1 c.6362G>A and @GENE$ @VARIANT$), 2F07 (CELSR1 c.8807C>T and @GENE$ c.1622C>T), 618F05 (CELSR1 c.8282C>T and SCRIB @VARIANT$).

PRICKLE4;22752

DVL3;20928

c.730C>G;tmVar:c|SUB|C|730|G;HGVS:c.730C>G;VariantGroup:12;CorrespondingGene:29964;RS#:141478229;CA#:3802865

c.3979G>A;tmVar:c|SUB|G|3979|A;HGVS:c.3979G>A;VariantGroup:31;CorrespondingGene:23513;RS#:201563528;CA#:4918429

0no label

M2, @GENE$: p.(E387K). M3, CYP1B1: @VARIANT$. M4, PITX2: p.(P179T). M5, @GENE$: @VARIANT$. Arrows show the index cases.

CYP1B1;68035

PITX2;55454

p.(E173*);tmVar:p|SUB|E|173|*;HGVS:p.E173*;VariantGroup:11;CorrespondingGene:1545

p.(A188T);tmVar:p|SUB|A|188|T;HGVS:p.A188T;VariantGroup:5;CorrespondingGene:5308;RS#:77144743;CA#:203139

0no label

Ebermann et al. described a USH2 patient with "digenic inheritance." a heterozygous truncating mutation in GPR98, and a truncating heterozygous mutation in PDZ domain-containing 7 (@GENE$), which is reported to be a cause of USH. Our USH1 patient (Case #4) had segregated MYO7A:p.Ala771Ser and PCDH15:@VARIANT$. Molecular analyses in mouse models have shown many interactions among the USH1 proteins. In particular, MYO7A directly binds to PCDH15 and both proteins are expressed in an overlapping pattern in hair bundles in a mouse model. @GENE$:c.158-1G>A, predicted to alter the splice donor site of intron 3, has been classified as pathogenic. MYO7A:@VARIANT$ is a non-truncating mutation, but was previously reported as disease-causing.

PDZD7;129509

PCDH15;23401

c.158-1G>A;tmVar:c|SUB|G|158-1|A;HGVS:c.158-1G>A;VariantGroup:18;CorrespondingGene:65217;RS#:876657418;CA#:10576348

p.Ala771Ser;tmVar:p|SUB|A|771|S;HGVS:p.A771S;VariantGroup:2;CorrespondingGene:4647;RS#:782384464;CA#:10576351

0no label

To investigate the role of @GENE$ variations along with GJB2 mutations for a possible combinatory allelic disease inheritance, we have screened patients with heterozygous GJB2 mutations for variants in Cx31 by sequencing. Analysis of the entire coding region of the Cx31 gene revealed the presence of two different missense mutations (N166S and A194T) occurring in compound heterozygosity along with the 235delC and 299delAT of @GENE$ in 3 simplex families (235delC/N166S, @VARIANT$/@VARIANT$ and 299delAT/A194T).

GJB3;7338

GJB2;2975

235delC;tmVar:c|DEL|235|C;HGVS:c.235delC;VariantGroup:1;CorrespondingGene:2706;RS#:80338943

A194T;tmVar:c|SUB|A|194|T;HGVS:c.194A>T;VariantGroup:4;CorrespondingGene:2707;RS#:117385606;CA#:118313

0no label

Compared to WT (wild-type) proteins, we found that the ability of GFP-CYP1B1 @VARIANT$ and GFP-@GENE$ E229K to immunoprecipitate HA-TEK E103D and HA-TEK Q214P, respectively, was significantly diminished. GFP-CYP1B1 R368H also exhibited relatively reduced ability to immunoprecipitate HA-@GENE$ @VARIANT$ (~70%).

CYP1B1;68035

TEK;397

A115P;tmVar:p|SUB|A|115|P;HGVS:p.A115P;VariantGroup:0;CorrespondingGene:1545;RS#:764338357;CA#:1620052

I148T;tmVar:p|SUB|I|148|T;HGVS:p.I148T;VariantGroup:5;CorrespondingGene:7010;RS#:35969327;CA#:5015918

0no label

Variants in all known WS candidate genes (EDN3, EDNRB, MITF, PAX3, @GENE$, SNAI2, and TYRO3) were searched and a novel rare heterozygous deletion mutation (@VARIANT$; p.Asn322fs) was identified in the @GENE$ gene in both patients. Moreover, heterozygous missense variants in SNAI3 (c.607C>T; @VARIANT$) and TYRO3 (c.1037T>A; p.Ile346Asn) gene was identified in the exome data of both patients.

SOX10;5055

MITF;4892

c.965delA;tmVar:c|DEL|965|A;HGVS:c.965delA;VariantGroup:4;CorrespondingGene:4286

p.Arg203Cys;tmVar:p|SUB|R|203|C;HGVS:p.R203C;VariantGroup:1;CorrespondingGene:333929;RS#:149676512;CA#:8229366

0no label

c, d) Sequence chromatograms indicating the wild-type, homozygous affected and heterozygous carrier forms of c) the C to T transition at position c.229 changing the @VARIANT$ of the @GENE$ protein (c.229C>T; p.R77C) and d) the @VARIANT$ (p.I80Gfs*13) in @GENE$. Mutation name is based on the full-length S100A3 (NM_002960) and S100A13 (NM_001024210) transcripts.

S100A3;2223

S100A13;7523

arginine residue to cysteine at position 77;tmVar:p|SUB|R|77|C;HGVS:p.R77C;VariantGroup:3;CorrespondingGene:6274;RS#:138355706;CA#:1116284

c.238-241delATTG;tmVar:c|DEL|238_241|ATTG;HGVS:c.238_241delATTG;VariantGroup:13;CorrespondingGene:6284

11

The DUSP6 gene [@VARIANT$; p.(Val114Leu)] was involved in all five disease-causing digenic combinations. Sanger sequencing showed that the @GENE$ variant [@VARIANT$; p.(Glu587Lys)] was only present in HH12 and absent in his asymptomatic mother (Figure 1). The variants located in the promoter region of @GENE$ were extracted, which revealed one common variant (c.-9 + 342A > G) in intron 1 with a MAF of 0.3 according to GnomAD.

SEMA7A;2678

PROKR2;16368

c.340G > T;tmVar:c|SUB|G|340|T;HGVS:c.340G>T;VariantGroup:5;CorrespondingGene:1848;RS#:2279574;CA#:6714072

c.1759G > A;tmVar:c|SUB|G|1759|A;HGVS:c.1759G>A;VariantGroup:7;CorrespondingGene:8482

0no label

Variants in all known WS candidate genes (@GENE$, EDNRB, MITF, PAX3, SOX10, SNAI2, and @GENE$) were searched and a novel rare heterozygous deletion mutation (c.965delA; @VARIANT$) was identified in the MITF gene in both patients. Moreover, heterozygous missense variants in SNAI3 (c.607C>T; p.Arg203Cys) and TYRO3 (@VARIANT$; p.Ile346Asn) gene was identified in the exome data of both patients.

EDN3;88

TYRO3;4585

p.Asn322fs;tmVar:p|FS|N|322||;HGVS:p.N322fsX;VariantGroup:3;CorrespondingGene:4286

c.1037T>A;tmVar:c|SUB|T|1037|A;HGVS:c.1037T>A;VariantGroup:2;CorrespondingGene:7301;RS#:12148316;CA#:7494886

0no label

These results suggest an important role of ephrin-B2 as an inducer of @GENE$ endocytosis with the transmembrane binding partner, pendrin, while its effect is weaker than that of ephrin-A1. Aberrant regulation of pathogenic forms of pendrin via EphA2 Some pathogenic variants of pendrin are not affected by EphA2/ephrin-B2 regulation. a, b Immunoprecipitation of EphA2 with mutated pendrin. myc-pendrin @VARIANT$, L445W, Q446R, G672E were not co-immunoprecipitated with EphA2. Densitometric quantifications are shown (b). Mean +- SEM; one-way ANOVA with Bonferroni post hoc analyses; *p < 0.05; (n = 3). c, d Immunoprecipitation of EphA2 with mutated pendrin. Immunocomplex of myc-pendrin L117F, @VARIANT$ and F355L was not affected. Densitometric quantifications are shown (d). Mean +- SEM; (n = 3). e, f Internalization of EphA2 and mutated pendrin triggered by @GENE$ stimulation.

EphA2;20929

ephrin-B2;3019

A372V;tmVar:p|SUB|A|372|V;HGVS:p.A372V;VariantGroup:11;CorrespondingGene:5172;RS#:121908364;CA#:253306

S166N;tmVar:p|SUB|S|166|N;HGVS:p.S166N;VariantGroup:22;CorrespondingGene:23985

0no label

A total of 2 novel variants, @VARIANT$ and @VARIANT$, were located in a myeloperoxidase-like domain, the catalytic site of the enzyme (Fig. S3B). A total of 4 @GENE$ variants were found in 2 patients and were compound heterozygotes for 2 different TSHR mutations. The TSHR variant p.R450H was a recurrent inactivating mutation and p.C176R and p.K618 were novel. p.C176R is located in the leucine-rich repeat region of the extracellular domain and responsible for high-affinity hormone binding and p.R528S and p.K618* are located in the cytoplasmic loops (Fig. S3C). Patients with GIS had a higher tendency to be affected with mutations than patients with TD [25/32 (78%) vs. 6/11 (54%), Fig. 2]. Variants in TG, TSHR, @GENE$, SLC5A5 and PROP1 genes were found exclusively in patients with GIS, and 1 variant in TRHR was found in patients with TD.

TSHR;315

DUOXA2;57037

p.S309P;tmVar:p|SUB|S|309|P;HGVS:p.S309P;VariantGroup:13;CorrespondingGene:2304;RS#:1162674885

p.S571R;tmVar:p|SUB|S|571|R;HGVS:p.S571R;VariantGroup:26;CorrespondingGene:79048;RS#:765990605

0no label

Variants in all known WS candidate genes (EDN3, EDNRB, MITF, PAX3, SOX10, SNAI2, and @GENE$) were searched and a novel rare heterozygous deletion mutation (c.965delA; @VARIANT$) was identified in the @GENE$ gene in both patients. Moreover, heterozygous missense variants in SNAI3 (@VARIANT$; p.Arg203Cys) and TYRO3 (c.1037T>A; p.Ile346Asn) gene was identified in the exome data of both patients.

TYRO3;4585

MITF;4892

p.Asn322fs;tmVar:p|FS|N|322||;HGVS:p.N322fsX;VariantGroup:3;CorrespondingGene:4286

c.607C>T;tmVar:c|SUB|C|607|T;HGVS:c.607C>T;VariantGroup:1;CorrespondingGene:333929;RS#:149676512;CA#:8229366

0no label

The proband, who had @GENE$ p.(Asn1075Ser), p.(@VARIANT$), and @GENE$ p.(@VARIANT$) variants, showed ten missing teeth, while her parents, who passed individual mutant alleles, had no missing teeth but microdontia and dysmorphology of specific teeth.

LRP6;1747

WNT10A;22525

Ser127Thr;tmVar:p|SUB|S|127|T;HGVS:p.S127T;VariantGroup:1;CorrespondingGene:4040;RS#:17848270;CA#:6455897

Glu167Gln;tmVar:p|SUB|E|167|Q;HGVS:p.E167Q;VariantGroup:5;CorrespondingGene:80326;RS#:148714379

11

Genetic evaluation revealed heterozygous variants in the related genes NRXN1 (c.2686C>T, @VARIANT$) and NRXN2 (@VARIANT$, p.Arg1059Gln), one inherited from the mother with family history of sudden infant death syndrome (SIDS) and one from the father with family history of febrile seizures. Although there are no previous reports with the digenic combination of NRXN1 and NRXN2 variants, patients with biallelic loss of NRXN1 in humans and double neurexin 1alpha/2alpha knockout mice have severe breathing abnormalities, corresponding to the respiratory phenotype of our patient. These observations and the known interaction between the NRXN1 and NRXN2 proteins lead us to hypothesize that digenic variants in @GENE$ and @GENE$ contributed to the phenotype of EIEE, arcuate nucleus hypoplasia, respiratory failure, and death.

NRXN1;21005

NRXN2;86984

p.Arg896Trp;tmVar:p|SUB|R|896|W;HGVS:p.R896W;VariantGroup:1;CorrespondingGene:9378;RS#:796052777;CA#:316143

c.3176G>A;tmVar:c|SUB|G|3176|A;HGVS:c.3176G>A;VariantGroup:2;CorrespondingGene:9379;RS#:777033569;CA#:6078001

11

Two unrelated KS patients had heterozygous NELF mutations and mutation in a second gene: NELF/KAL1 (c.757G>A; p.Ala253Thr of NELF and @VARIANT$; @VARIANT$ of KAL1) and @GENE$/@GENE$ (c. 1160-13C>T of NELF and c.824G>A; p.Trp275X of TACR3).

NELF;10648

TACR3;824

c.488_490delGTT;tmVar:p|DEL|488_490|V;HGVS:p.488_490delV;VariantGroup:8;CorrespondingGene:26012

p.Cys163del;tmVar:p|DEL|163|C;HGVS:p.163delC;VariantGroup:10;CorrespondingGene:3730

0no label

Cardiac Phenotype: A @GENE$/NFATC1 Genetic Interaction The cardiac phenotype in the indexed-family is divided into two: a mild VSD not requiring any intervention and a severe TOF-like phenotype that required major intervention (Figure 1). We sought that differential variants inherited from the father would contribute to this differential expressivity of the ARS phenotype within the three affected children in this family: two with a cardiac phenotype and the third with only glaucoma. Interestingly, we unravel two novel missense mutations in @GENE$ (@VARIANT$) and NFATC1 (@VARIANT$) that are predicted to be damaging (Table 4).

FOXC1;20373

OBSCN;70869

p.C1880Y;tmVar:p|SUB|C|1880|Y;HGVS:p.C1880Y;VariantGroup:129;CorrespondingGene:84033

p.R222Q;tmVar:p|SUB|R|222|Q;HGVS:p.R222Q;VariantGroup:10;CorrespondingGene:4772;RS#:1390597692

0no label

The proband's mother and aunt, also manifesting with PXE-like skin changes, were heterozygous carriers of a missense mutation (@VARIANT$) in @GENE$ and a null mutation (@VARIANT$) in the @GENE$ gene, suggesting digenic nature of their skin findings.

GGCX;639

ABCC6;55559

p.V255M;tmVar:p|SUB|V|255|M;HGVS:p.V255M;VariantGroup:1;CorrespondingGene:2677;RS#:121909683;CA#:214957

p.R1141X;tmVar:p|SUB|R|1141|X;HGVS:p.R1141X;VariantGroup:6;CorrespondingGene:368;RS#:72653706;CA#:129115

11

The @GENE$ and SETDB1 variants were excluded based on their frequencies in normal population cohorts. Sanger sequencing of Family 1 showed that both @VARIANT$ in S100A3 (c.229C>T, missense causing a p.R77C mutation) and a 4 bp deletion in @GENE$ (@VARIANT$ causing a frameshift p.I80Gfs*13) segregated completely with ILD in Family 1 based upon recessive inheritance (figure 2c and d), were in total linkage disequilibrium, and were present in a cis conformation.

ISG20L2;12814

S100A13;7523

rs138355706;tmVar:rs138355706;VariantGroup:3;CorrespondingGene:6274;RS#:138355706

c.238-241delATTG;tmVar:c|DEL|238_241|ATTG;HGVS:c.238_241delATTG;VariantGroup:13;CorrespondingGene:6284

0no label

Moreover, mutations in residues close to @VARIANT$ and A194 identified in the families reported here, namely, M163L, R165W, F191L, and @VARIANT$ in Cx26 as well as F193C, S198F and G199R in Cx32, have been reported previously in patients with hearing impairment. Interestingly, mutations identified in patients with the skin disease erythrokeratoderma variabilis (EKV) were located within all the protein domains of the Cx31 gene except for the EC2 and TM4 domains, which are main domains for deafness mutations. This correlation between location of mutations and phenotypes, together with the identification of pathological mutations associated with hearing loss in the same region of the EC2 and TM4 domains in these three connexin genes (@GENE$, Cx31, and Cx32) suggested that the EC2 and TM4 domains are important to the function of the Cx31 protein in the inner ear and plays a vital role in forming connexons in the cells of the inner ear. In the present study, we have shown that the missense N166S and A194T mutations in @GENE$ acts in a recessive manner in three unrelated Chinese patients.

Cx26;2975

GJB3;7338

N166;tmVar:p|Allele|N|166;VariantGroup:0;CorrespondingGene:2707;RS#:121908851

A197S;tmVar:p|SUB|A|197|S;HGVS:p.A197S;VariantGroup:3;CorrespondingGene:2706;RS#:777236559

0no label

DISCUSSION In this study, we describe identification and characterization of abnormalities in patients with homozygous mutations in two genes, a novel mutation in SEC23A, @VARIANT$ and a previously identified mutation in MAN1B1, @VARIANT$. The affected patients presented with moderate global developmental delay, tall stature, obesity, macrocephaly, mild dysmorphic features, hypertelorism, maloccluded teeth, intellectual disability, and flat feet. We found that mutations in the two genes segregated in the family and that the unaffected parents were healthy and carried heterozygous mutations in both SEC23A and @GENE$, consistent with an autosomal-recessive mode of inheritance. We also identified heterozygous mutation in @GENE$ in an unaffected sibling of tall stature and normal intelligence.

MAN1B1;5230

SEC23A;4642

1200G>C;tmVar:c|SUB|G|1200|C;HGVS:c.1200G>C;VariantGroup:0;CorrespondingGene:10484;RS#:866845715;CA#:259543384

1000C>T;tmVar:c|SUB|C|1000|T;HGVS:c.1000C>T;VariantGroup:4;CorrespondingGene:11253;RS#:387906886;CA#:129197

0no label

RESULTS Mutations at the gap junction proteins @GENE$ and @GENE$ can interact to cause non-syndromic deafness In total, 108 probands screened for mutations in the Cx26 gene were found to carry a single recessive mutant allele. In those samples, no mutation was detected on the second allele either in Cx26-exon-1/splice sites or in GJB6. To investigate the role of GJB3 variations along with GJB2 mutations for a possible combinatory allelic disease inheritance, we have screened patients with heterozygous GJB2 mutations for variants in Cx31 by sequencing. Analysis of the entire coding region of the Cx31 gene revealed the presence of two different missense mutations (N166S and A194T) occurring in compound heterozygosity along with the 235delC and 299delAT of GJB2 in 3 simplex families (235delC/N166S, 235delC/A194T and 299delAT/A194T). In family A, a profoundly hearing impaired proband was found to be heterozygous for a novel @VARIANT$ of GJB3, resulting in an asparagine into serine substitution in codon 166 (N166S) and for the @VARIANT$ of GJB2 (Fig. 1b, d).

Cx26;2975

Cx31;7338

A to G transition at nucleotide position 497;tmVar:c|SUB|A|497|G;HGVS:c.497A>G;VariantGroup:0;CorrespondingGene:2707;RS#:121908851;CA#:118311

235delC;tmVar:c|DEL|235|C;HGVS:c.235delC;VariantGroup:1;CorrespondingGene:2706;RS#:80338943

11

Variants in all known WS candidate genes (EDN3, EDNRB, MITF, PAX3, SOX10, @GENE$, and TYRO3) were searched and a novel rare heterozygous deletion mutation (c.965delA; @VARIANT$) was identified in the MITF gene in both patients. Moreover, heterozygous missense variants in SNAI3 (c.607C>T; p.Arg203Cys) and @GENE$ (c.1037T>A; @VARIANT$) gene was identified in the exome data of both patients.

SNAI2;31127

TYRO3;4585

p.Asn322fs;tmVar:p|FS|N|322||;HGVS:p.N322fsX;VariantGroup:3;CorrespondingGene:4286

p.Ile346Asn;tmVar:p|SUB|I|346|N;HGVS:p.I346N;VariantGroup:2;CorrespondingGene:7301;RS#:12148316;CA#:7494886

0no label

Two unrelated KS patients had heterozygous NELF mutations and mutation in a second gene: NELF/KAL1 (c.757G>A; p.Ala253Thr of NELF and @VARIANT$; @VARIANT$ of @GENE$) and NELF/TACR3 (c. 1160-13C>T of @GENE$ and c.824G>A; p.Trp275X of TACR3).

KAL1;55445

NELF;10648

c.488_490delGTT;tmVar:p|DEL|488_490|V;HGVS:p.488_490delV;VariantGroup:8;CorrespondingGene:26012

p.Cys163del;tmVar:p|DEL|163|C;HGVS:p.163delC;VariantGroup:10;CorrespondingGene:3730

0no label

Similarly, patients 8 and 10 both had a combination of a known truncating mutation (@VARIANT$) and a known inactivating mutation (@VARIANT$ or p.R885Q); one exhibited permanent CH and one showed transient hypothyroidism. Furthermore, patient 7 had exactly the same mutations as patient 8, and her prognosis was unknown. Unlike patient 8, who had a goiter, patient 7's thyroid size was normal. Moreover, numbers of detected variants differed among patients who shared the same phenotypes. 4. Discussion Thyroid hormone biosynthesis defects are common causes of CH. Mutations in DH-associated genes, including @GENE$, @GENE$, DUOX2, DUOXA2, SLC26A4, SCL5A5, and IYD, have been detected in numerous cases.

TPO;461

TG;2430

p.K530X;tmVar:p|SUB|K|530|X;HGVS:p.K530X;VariantGroup:6;CorrespondingGene:50506;RS#:180671269;CA#:7538552

p.R110Q;tmVar:p|SUB|R|110|Q;HGVS:p.R110Q;VariantGroup:29;CorrespondingGene:7173;RS#:750143029;CA#:1511376

0no label

Results Cosegregating deleterious variants (GRCH37/hg19) in @GENE$ (NM_001127222.1: c.7261_7262delinsGT, @VARIANT$), REEP4 (NM_025232.3: c.109C>T, p.Arg37Trp), TOR2A (NM_130459.3: c.568C>T, @VARIANT$), and @GENE$ (NM_005173.3: c.1966C>T, p.Arg656Cys) were identified in four independent multigenerational pedigrees.

CACNA1A;56383

ATP2A3;69131

p.Pro2421Val;tmVar:p|SUB|P|2421|V;HGVS:p.P2421V;VariantGroup:3;CorrespondingGene:80346

p.Arg190Cys;tmVar:p|SUB|R|190|C;HGVS:p.R190C;VariantGroup:12;CorrespondingGene:27433;RS#:376074923;CA#:5250615

0no label

Five anencephaly cases carried rare or novel CELSR1 missense variants, three of whom carried additional rare potentially damaging PCP variants: 01F377 (CELSR1 c.6362G>A and PRICKLE4 @VARIANT$), 2F07 (CELSR1 c.8807C>T and DVL3 c.1622C>T), 618F05 (CELSR1 @VARIANT$ and SCRIB c.3979G>A). One patient (f93-80) had a novel PTK7 missense variant (c.655A>G) with a rare CELSR2 missense variant (c.1892C>T). Three patients carried missense variants both in @GENE$ and other PCP-associated genes: 01F552 (FZD6 c.1531C>T and CELSR2 c.3800A>G), 335F07 (FZD6 c.544G>A and 2 FAT4 missense variants c.5792A>G; c.10384A>G), and 465F99 (rare FZD1 missense variant c.211C>T and a novel @GENE$ missense variant c.10147G>A).

FZD;8321;8323

FAT4;14377

c.730C>G;tmVar:c|SUB|C|730|G;HGVS:c.730C>G;VariantGroup:12;CorrespondingGene:29964;RS#:141478229;CA#:3802865

c.8282C>T;tmVar:c|SUB|C|8282|T;HGVS:c.8282C>T;VariantGroup:4;CorrespondingGene:9620;RS#:144039991;CA#:10292903

0no label

Altogether, the results suggest that @GENE$ @VARIANT$ is a loss-of-function mutation in Drosophila. KAT2B F307S but not @GENE$ @VARIANT$ causes cardiac defects in Drosophila Since the presence of SRNS and heart defects in family A was the main phenotypic difference from the other families, we looked more specifically into the cardiac and renal system of the fly.

KAT2B;20834

ADD3;40893

F307S;tmVar:p|SUB|F|307|S;HGVS:p.F307S;VariantGroup:1;CorrespondingGene:8850

E659Q;tmVar:p|SUB|E|659|Q;HGVS:p.E659Q;VariantGroup:4;CorrespondingGene:120;RS#:753083630;CA#:5686787

0no label

Both homozygous and compound heterozygous variants in the @GENE$ gene have been described as causative for juvenile ALS. The @VARIANT$ nonsense variant was first detected in compound heterozygous form in a family with two affected siblings suffering from infantile ascending spastic paralysis with bulbar involvement. The ages of onset of the patients with the ALS2 variants reported in this study were later than juvenile ALS onset, which generally manifests before 25 years of age. Previous studies suggested that heterozygous variants in the ALS2 may be causative for adult-onset sALS. @GENE$ encodes three protein isoforms that have been described as nuclear-matrix and DNA/RNA binding proteins involved in transcription and stabilization of mRNA. In the present study, two novel heterozygous variants (P11S, S275N) were detected. The P11S variant affects the b isoform of the MATR3 protein (NM_001194956 and NP_001181885), contributing to splicing alteration of other isoforms. Further evidence is required to elucidate the mechanism of pathogenicity of these alterations. We discovered several variants in ALS candidate and risk genes. In a patient with LMN-dominant ALS with slow progression, we found two novel variants (@VARIANT$ and G4290R) in the DYNC1H1 gene.

ALS2;23264

MATR3;7830

G1177X;tmVar:p|SUB|G|1177|X;HGVS:p.G1177X;VariantGroup:0;CorrespondingGene:57679;RS#:386134180;CA#:356568

T2583I;tmVar:p|SUB|T|2583|I;HGVS:p.T2583I;VariantGroup:31;CorrespondingGene:1778

0no label

The @VARIANT$ mutation would be predicted to generate a nonfunctional @GENE$ enzyme, and its digenic inheritance alongside the homozygous DUOX2 @VARIANT$ will likely result in complete absence of functional DUOX isoenzyme in our patients. It has been speculated that DUOX1 upregulation in the context of DUOX2 loss of function may at least partially compensate for defective H2O2 production. In support of this notion, the majority of reported biallelic @GENE$ mutations, which are known to truncate the protein before the H2O2-generating domains, cause transient or mild permanent CH, despite presumably abrogating DUOX2 activity completely (Table 1).

DUOX1;68136

DUOX2;9689

c.1823-1G>C;tmVar:c|SUB|G|1823-1|C;HGVS:c.1823-1G>C;VariantGroup:17;CorrespondingGene:53905

p.R434*;tmVar:p|SUB|R|434|*;HGVS:p.R434*;VariantGroup:0;CorrespondingGene:50506;RS#:119472026

0no label

CONCLUSIONS We firstly identified the novel digenic heterozygous mutations by WES, KCNH2 @VARIANT$ and @GENE$ @VARIANT$, which resulted in LQTS with repeat syncope, torsades de pointes, ventricular fibrillation, and sinoatrial node dysfunction. KCNH2 p.307_308del may affect the function of @GENE$ channel in cardiomyocytes by inducing a regional double helix of the amino acids misfolded and largest hydrophobic domain disorganized.

SCN5A;22738

Kv11.1;201

p.307_308del;tmVar:p|DEL|307_308|;HGVS:p.307_308del;VariantGroup:16;CorrespondingGene:3757

p.R1865H;tmVar:p|SUB|R|1865|H;HGVS:p.R1865H;VariantGroup:1;CorrespondingGene:6331;RS#:370694515;CA#:64651

0no label

Two unrelated KS patients had heterozygous NELF mutations and mutation in a second gene: NELF/@GENE$ (c.757G>A; p.Ala253Thr of NELF and c.488_490delGTT; p.Cys163del of KAL1) and NELF/TACR3 (@VARIANT$ of NELF and c.824G>A; @VARIANT$ of @GENE$).

KAL1;55445

TACR3;824

c. 1160-13C>T;tmVar:c|SUB|C|1160-13|T;HGVS:c.1160-13C>T;VariantGroup:5;CorrespondingGene:26012;RS#:781275840;CA#:5370137

p.Trp275X;tmVar:p|SUB|W|275|X;HGVS:p.W275X;VariantGroup:1;CorrespondingGene:6870;RS#:144292455;CA#:144871

0no label

Moreover, patients carrying a @GENE$ @VARIANT$ mutation have a significantly reduced extracellular matrix (ECM) in cardiomyocytes. These findings support the importance of LAMA4 as a structural and signalling molecule in cardiomyocytes, and may indicate the modifier role that missense variations in LAMA4 play in the disease. Digenic heterozygosity has been described in some DCM cases and is often associated with a severe presentation of DCM. Moller et al. reported an index case with digenic variants in MYH7 (@VARIANT$) and @GENE$ (R326Q), both encoding sarcomeric proteins that are likely to affect its structure when mutated.

LAMA4;37604

MYBPC3;215

Pro943Leu;tmVar:p|SUB|P|943|L;HGVS:p.P943L;VariantGroup:5;CorrespondingGene:3910;RS#:387907365;CA#:143749

L1038P;tmVar:p|SUB|L|1038|P;HGVS:p.L1038P;VariantGroup:8;CorrespondingGene:4625;RS#:551897533;CA#:257817954

0no label

Notably, the patients carrying the @VARIANT$ and p.I400V mutations, and three patients carrying the @VARIANT$ mutation also carry, in heterozygous state, p.Y217D, p.R268C (two patients), p.H70fsX5, and p.G687N pathogenic mutations in KAL1, PROKR2, @GENE$, and @GENE$, respectively (Table 1), which further substantiates the digenic/oligogenic mode of inheritance of KS.

PROK2;9268

FGFR1;69065

p.T688A;tmVar:p|SUB|T|688|A;HGVS:p.T688A;VariantGroup:0;CorrespondingGene:2260;RS#:876661335

p.V435I;tmVar:p|SUB|V|435|I;HGVS:p.V435I;VariantGroup:1;CorrespondingGene:10371;RS#:147436181;CA#:130481

0no label

One fetus with anencephaly (735F97) carried a rare missense mutation (c.2852C>A; @VARIANT$; rs147472391) in glycine decarboxylase (GLDC), which was previously reported as one of the causative mutated alleles in a compound heterozygous patient with the autosomal recessive disorder, non-ketotic hyperglycinemia (NKH, OMIM: 605899).30 @GENE$ encodes a component of the glycine cleavage system (GCS) in mitochondrial folate one-carbon metabolism, which has previously been implicated in both mouse and human NTDs.10, 11, 31, 32 Another individual with anencephaly (706F07) was heterozygous for a missense variant (c.200C>T; p.Thr67Ile; @VARIANT$) in the @GENE$ (CTH) gene, which causes cystathioninuria in a recessive form.33 Unreported and rare variants After filtering out intron, 3' and 5' UTR and synonymous variants, a total of 397 rare (MAF < 1%, including novel) variants were identified in 89/90 NTD cases from 110/191 of the panel of NTD candidate genes.

GLDC;141

cystathionine gamma-lyase;1432

p.Ser951Tyr;tmVar:p|SUB|S|951|Y;HGVS:p.S951Y;VariantGroup:20;CorrespondingGene:2731;RS#:147472391;CA#:4980035

rs28941785;tmVar:rs28941785;VariantGroup:30;CorrespondingGene:1491;RS#:28941785

0no label

The nucleotide sequence showed a G to C transition at nucleotide 769 (c.769G>C) of the coding sequence in exon 7 of @GENE$, which results in the substitution of Gly at residue 257 to Arg. Additionally, the nucleotide sequence showed a monoallelic C to T transition at nucleotide 511 (c.511C>T) of the coding sequence in exon 3 of WNT10A, which results in the substitution of Arg at residue 171 to Cys. DNA sequencing of the parents' genome revealed that both mutant alleles were from their mother (Fig. 2A), who carried a heterozygous EDA mutation (c.769G>C) and a heterozygous @GENE$ @VARIANT$ mutation, and showed absence of only the left upper lateral incisor without other clinical abnormalities. No mutations in these genes were found in the father. Sequence analyses of EDA and WNT10A genes. (A) The EDA mutation @VARIANT$ and WNT10A mutation c.511C>T were found in patient N1, who inherited the mutant allele from his mother.

EDA;1896

WNT10A;22525

c.511C>T;tmVar:c|SUB|C|511|T;HGVS:c.511C>T;VariantGroup:3;CorrespondingGene:80326;RS#:116998555;CA#:2113955

c.769G>C;tmVar:c|SUB|G|769|C;HGVS:c.769G>C;VariantGroup:0;CorrespondingGene:1896;RS#:1057517882;CA#:16043329

0no label

This de novo variant may modify the effect of the truncating variant in @GENE$ by repressing @GENE$/TGF-beta signalling. In patient AVM359, one heterozygous VUS (@VARIANT$ [p.Arg197Trp]) in ENG inherited from the mother and one likely pathogenic de novo heterozygous variant (@VARIANT$ [p.Cys531Tyr]) in SCUBE2 were identified (online supplementary table S2).

ENG;92

BMP;55955

c.589C>T;tmVar:c|SUB|C|589|T;HGVS:c.589C>T;VariantGroup:2;CorrespondingGene:83394;RS#:2229778;CA#:2061380

c.1592G>A;tmVar:c|SUB|G|1592|A;HGVS:c.1592G>A;VariantGroup:5;CorrespondingGene:1956;RS#:1212415588

0no label

Here, we have demonstrated that the @GENE$ @VARIANT$ mutation has a more detrimental effect on the phenotype in this pedigree. It could be argued that the TNFRSF13B/@GENE$ @VARIANT$ mutation has a modifying effect on the phenotype and is relatively benign in this family.

TCF3;2408

TACI;49320

T168fsX191;tmVar:p|FS|T|168||191;HGVS:p.T168fsX191;VariantGroup:1;CorrespondingGene:6929

C104R;tmVar:p|SUB|C|104|R;HGVS:p.C104R;VariantGroup:2;CorrespondingGene:23495;RS#:34557412;CA#:117387

11

Two affected (II-3 and III-9) individuals were selected for WES. +/+, wild-type; +/-, heterozygous for REEP4 c.109C>T. (b) Electropherograms of unaffected family member (II-2) and subject with @GENE$ (II-3). (c) Multiple sequence alignment shows evolutionary conservation of @VARIANT$ among vertebrates @GENE$ missense variant A TOR2A nonsynonymous SNV (@VARIANT$ [NM_130459.3], p.Arg190Cys [NP_569726.2]) was identified in three subjects with BSP and three asymptomatic members from a four generation pedigree (Figure 5; Tables 1, 5, 8 and S2; Data S1).

BSP+;3644

TOR2A;25260

Arg37;tmVar:p|Allele|R|37;VariantGroup:10;CorrespondingGene:80346;RS#:780399718

c.568C>T;tmVar:c|SUB|C|568|T;HGVS:c.568C>T;VariantGroup:12;CorrespondingGene:27433;RS#:376074923;CA#:5250615

0no label

Additionally, the nucleotide sequence showed a monoallelic @VARIANT$ (c.511C>T) of the coding sequence in exon 3 of WNT10A, which results in the substitution of Arg at residue 171 to Cys. DNA sequencing of the parents' genome revealed that both mutant alleles were from their mother (Fig. 2A), who carried a heterozygous @GENE$ mutation (@VARIANT$) and a heterozygous WNT10A c.511C>T mutation, and showed absence of only the left upper lateral incisor without other clinical abnormalities. No mutations in these genes were found in the father. Sequence analyses of EDA and @GENE$ genes.

EDA;1896

WNT10A;22525

C to T transition at nucleotide 511;tmVar:c|SUB|C|511|T;HGVS:c.511C>T;VariantGroup:3;CorrespondingGene:80326;RS#:116998555;CA#:2113955

c.769G>C;tmVar:c|SUB|G|769|C;HGVS:c.769G>C;VariantGroup:0;CorrespondingGene:1896;RS#:1057517882;CA#:16043329

0no label

In this study, we speculated that, during the repolarization phase, the inadequate inward current caused by the detrimental CACNA1C-Q1916R mutation might be partly compensated by the persistent inward tail INa produced by the @GENE$-@VARIANT$ channel. That may be how SCN5A-R1193Q plays a protective role against the detrimental phenotype induced by the @GENE$-@VARIANT$ mutation.

SCN5A;22738

CACNA1C;55484

R1193Q;tmVar:p|SUB|R|1193|Q;HGVS:p.R1193Q;VariantGroup:7;CorrespondingGene:6331;RS#:41261344;CA#:17287

Q1916R;tmVar:p|SUB|Q|1916|R;HGVS:p.Q1916R;VariantGroup:4;CorrespondingGene:775;RS#:186867242;CA#:6389963

0no label

Two unrelated KS patients had heterozygous NELF mutations and mutation in a second gene: NELF/KAL1 (c.757G>A; p.Ala253Thr of @GENE$ and @VARIANT$; @VARIANT$ of @GENE$) and NELF/TACR3 (c. 1160-13C>T of NELF and c.824G>A; p.Trp275X of TACR3).

NELF;10648

KAL1;55445

c.488_490delGTT;tmVar:p|DEL|488_490|V;HGVS:p.488_490delV;VariantGroup:8;CorrespondingGene:26012

p.Cys163del;tmVar:p|DEL|163|C;HGVS:p.163delC;VariantGroup:10;CorrespondingGene:3730

0no label