1-179557079-C-T
Variant summary
Our verdict is Likely benign. Variant got -1 ACMG points: 5P and 6B. PM1PM5PP5BP4BS1_SupportingBS2
The NM_014625.4(NPHS2):c.686G>A(p.Arg229Gln) variant causes a missense change involving the alteration of a non-conserved nucleotide. The variant allele was found at a frequency of 0.0348 in 1,613,872 control chromosomes in the GnomAD database, including 1,141 homozygotes. In-silico tool predicts a benign outcome for this variant. Variant has been reported in ClinVar as Conflicting classifications of pathogenicity (no stars). Another variant affecting the same amino acid position, but resulting in a different missense (i.e. R229L) has been classified as Likely pathogenic.
Frequency
Consequence
NM_014625.4 missense
Scores
Clinical Significance
Conservation
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ACMG classification
Verdict is Likely_benign. Variant got -1 ACMG points.
Transcripts
RefSeq
Ensembl
Frequencies
GnomAD3 genomes 0.0277 AC: 4222AN: 152170Hom.: 85 Cov.: 32
GnomAD3 exomes AF: 0.0304 AC: 7639AN: 250894Hom.: 170 AF XY: 0.0313 AC XY: 4248AN XY: 135594
GnomAD4 exome AF: 0.0355 AC: 51908AN: 1461584Hom.: 1056 Cov.: 32 AF XY: 0.0356 AC XY: 25877AN XY: 727076
GnomAD4 genome 0.0277 AC: 4222AN: 152288Hom.: 85 Cov.: 32 AF XY: 0.0284 AC XY: 2112AN XY: 74458
ClinVar
Submissions by phenotype
Nephrotic syndrome, type 2 Pathogenic:6Uncertain:5
The NPHS2 c.686G>A variant is classified as LIKELY PATHOGENIC (PS4_Moderate, PM3, PS3, PM5_Supporting) The NPHS2 c.686G>A variant is a single nucleotide change in exon 5/8 of the NPHS2 gene, which is predicted to change the amino acid arginine at position 229 in the protein to glutamine. The variant has been reported frequently in a compound heterozygous state in probands with a clinical presentation of nephrotic syndrome and focal segmental glomerulosclerosis (PS4_Moderate). The variant is common in population databases (PM2 not met) and has been reported as a disease-associated polymorphism, having been frequently reported in trans with pathogenic variants, including the p.Ala297Val variant also detected in this patient (Tory et al. 2014 PMID:24509478) (PM3). Transfection studies in human podocyte cell lines showed that when the p.Ala297Val variant was co-expressed with p.Arg229Gln (also in this patient), podocin was retained in the cytoplasm, rather than localising properly to the plasma membrane (Tory et al. 2014 PMID:24509478) (PS3). Computational structural modelling showed the p.Ala297Val variant in trans with p.Arg229Gln led to altered dimerisation. They predicted this most likely contributed to the retention of podocin within cytoplasmic compartments. This variant is a missense change at an amino acid residue where different missense changes have been seen before (HGMD CM077322 - c.686G>T; p.R229L) (HGMD CM077322 - c.686G>T; p.R229L) (PM5_supporting). The variant has been reported in dbSNP (rs61747728) and in the HGMD database: CM023107. It has been reported with as pathogenic by other diagnostic laboratories (ClinVar Variation ID: 5370). -
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This variant was observed as part of a predisposition screen in an ostensibly healthy population. A literature search was performed for the gene, cDNA change, and amino acid change (where applicable). Publications were found based on this search. However, the evidence from the literature, in combination with allele frequency data from public databases where available, was not sufficient to rule this variant in or out of causing disease. Therefore, this variant is classified as a variant of unknown significance. -
The pathogenicity of R229Q is dependent on the variant observed on the other chromosome. In homozygous state, R229Q is not disease-causing. -
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PM1:Located in well-established functional domain PP3:Multiple lines of computational evidence support a deleterious effect on the gene or gene product -
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This missense variant (c.686G>A, p.Arg229Gln) has been observed in population databases (gnomAD) and is reported in the literature (PMID 24509478, 24715228, 24072153, 19145239, 18823551 12464671). Variant prediction programs do not show agreement regarding the effect of this variant, although functional studies suggest alteration in dimerization and membrane localization when in trans with another NPHS2 variant . This variant was identified in an affected patient. -
Based on the classification scheme VCGS_Germline_v1.3.4, this variant is classified as Likely pathogenic. Following criteria are met: 0102 - Loss of function is a known mechanism of disease in this gene and is associated with nephrotic syndrome, type 2 (MIM#600995). (I) 0106 - This gene is associated with autosomal recessive disease. (I) 0200 - Variant is predicted to result in a missense amino acid change from arginine to glutamine. (I) 0251 - This variant is heterozygous. (I) 0307 - Variant is present in gnomAD (v2) at a frequency >=0.05 (8166 heterozygotes, 186 homozygotes). (SB) 0502 - Missense variant with conflicting in silico predictions and uninformative conservation. (I) 0600 - Variant is located in the annotated SPFH domain (NCBI). (I) 0705 - No comparable missense variants have previous evidence for pathogenicity. (I) 0801 - This variant has strong previous evidence of pathogenicity in unrelated individuals. This variant is commonly reported to have variant-dependent pathogenicity. It has been reported in many patients with nephrotic syndrome or adult-onset focal segmental glomerulosclerosis as compound heterozygous with specific missense variants. However, incomplete penetrance has also been reported; the phenotype may be less severe, where age at diagnosis is delayed and disease progression is slower. This variant is not purported to cause disease when in homozygous state (PMIDs: 24509478, 30348286, 30260545, 32467597, ClinVar). (SP) 1002 - This variant has moderate functional evidence supporting abnormal protein function. Functional analysis showed NPHS2-R229Q had reduced binding with NPHS1 (PMID: 12464671). Co-expression of NPHS2-R229Q and NPHS2 with different pathogenic missense variants showed the proteins were mislocalised (PMID: 24509478). (SP) 1208 - Inheritance information for this variant is not currently available in this individual. (I) Legend: (SP) - Supporting pathogenic, (I) - Information, (SB) - Supporting benign -
not provided Pathogenic:2Uncertain:3
In silico analysis supports that this missense variant does not alter protein structure/function; This variant is associated with the following publications: (PMID: 30777632, 14871423, 23800802, 12464671, 24509478, 28780565, 30241959, 28712774, 21415313, 18499321, 21722858, 23349334, 32691731, 33144682, 24715228, 23515051, 33193607, 20947785, 20798252, 30013592, 24856380, 19145239, 18726620, 26211502, 25903641, 27701157, 27004616, 27312921, 16354237, 16898497, 29869118, 27885584, 30450462, 30808327, 30260545, 29127259, 24519673, 31949506, 30586318, 33838161, 35253369, 22228437, 35064937, 34405919, 31027891, 38170106, 26138234, 30348286, 37204080, 35935761, 35325889, 33532864, 31308072, 37372416, 38765578, 15327385, 24072153, 38336226, 17371932, 15817495, 34853150, 18216321, 26420286, 26413278) -
While the frequency of this variant in the general population is higher than would generally be expected for pathogenic variants in this gene, it is also statistically more frequent in affected individuals than in the general population and/or healthy controls (PMID: 21415313, 26420286, 26413278, 20798252, 20947785 19145239, 18216321; (Genome Aggregation Database (gnomAD), Cambridge, MA (URL: http://gnomad.broadinstitute.org)). According to published literature (PMID: 24509478, 30260545) this variant only leads to disease when it is found in trans with a second pathogenic variant located within the codon range of 270-351 of the NPHS2 gene. This variant segregates with disease in multiple families (PMID: 12464671, 26413278, 26420286). Assessment of experimental evidence suggests this variant results in abnormal protein function. Experiments demonstrate this variant results in reduced binding of podocin to nephrin (PMID: 12464671, 24509478). In multiple individuals, this variant has been seen with a single recessive pathogenic variant in the same gene, suggesting this variant may also be pathogenic. -
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The NPHS2 p.Arg229Gln variant has been reported in multiple cases of steroid-resistant nephrotic syndrome (SRNS) (McCarthy_2013_PMID:23349334; Karle_2002_PMID:11805166; Ali_2017_PMID:28529802), however this variant is also found in control databases in 8538 of 282294 chromosomes (186 homozygous) at a frequency of 0.030245 increasing the likelihood this could be a low frequency benign variant (Genome Aggregation Database March 6, 2019, v2.1.1). The variant was observed in the following populations: European (Finnish) in 1715 of 25074 chromosomes (freq: 0.0684), Ashkenazi Jewish in 535 of 10354 chromosomes (freq: 0.05167), European (non-Finnish) in 4639 of 128830 chromosomes (freq: 0.03601), Other in 223 of 7196 chromosomes (freq: 0.03099), South Asian in 825 of 30590 chromosomes (freq: 0.02697), Latino in 458 of 35342 chromosomes (freq: 0.01296), African in 141 of 24966 chromosomes (freq: 0.005648), and East Asian in 2 of 19942 chromosomes (freq: 0.0001). The variant was also identified in dbSNP (ID: rs61747728), LOVD 3.0 (classified as a VUS and likely pathogenic) and ClinVar (classified as pathogenic by Athena Diagnostics and the Gharavi Laboratory at Columbia University, as likely pathogenic by Counsyl and as a VUS by Blueprint Genetics and Integrated Genetics). The p.Arg229 residue is conserved in mammals but not in more distantly related organisms however computational analyses (PolyPhen-2, SIFT, AlignGVGD, BLOSUM, MutationTaster) do not suggest a high likelihood of impact to the protein; this information is not predictive enough to rule out pathogenicity. The variant occurs outside of the splicing consensus sequence and in silico or computational prediction software programs (SpliceSiteFinder, MaxEntScan, NNSPLICE, GeneSplicer) do not predict a difference in splicing. A meta-analysis of the role of the p.R229Q variant in disease found significantly higher rates of SRNS in individuals homozygous for the variant compared to individuals homozygous for the reference allele (Lu_2014_PMID:24715228). However, no association between the p.R229Q variant was found in relation to late-onset focal segmental glomerulosclerosis (FSGS) or childhood-onset nephrotic syndrome in case-control studies (Hashemi_2015_PMID:25599733; McKenzie_2007_PMID:17942957). Kerti et al. (2013) identified a 37-year-old proband with proteinuria, FSGS and end-stage renal disease who was homozygous for the p.R229Q variant and also carried a truncating variant in the PAX2 gene. The proband's unaffected father and brother were also homozygous for the p.R229Q variant suggesting that it is a modifer of disease and not causal (Kerti_2013_PMID:23800802). A comprehensive study also suggests the p.R229Q variant is only pathogenic when found in trans with known pathogenic NPHS2 3' (C-terminal) variants, such as p.A284V, p.A288T, p.R291W, p.A297V or p.E310K. Functional studies of the p.R229Q variant with these variants demonstrated abnormal localization (Tory_2014_PMID:24509478). This suggests that the pathogenicity of the p.R229Q variant is dependent on which variant it is found in trans with. In summary, this variant is classified as a variant of uncertain significance. -
This sequence change replaces arginine, which is basic and polar, with glutamine, which is neutral and polar, at codon 229 of the NPHS2 protein (p.Arg229Gln). This variant is present in population databases (rs61747728, gnomAD 7%), including at least one homozygous and/or hemizygous individual. This variant has been observed in many individuals with adolescent/early adult onset nephrotic syndrome or focal segmental glomerulosclerosis (FSGS) and been observed to segregate with disease (PMID: 12464671, 19145239, 20947785, internal data). However, it appears to be associated with disease only when in trans with certain rare NPHS2 variants (e.g., p.Phe344Leufs*4, p.Ala284Val). When present in homozygosity or in combination with certain other rare NPHS2 variants (e.g., p.Arg138*, p.Val290Met), it is not associated with disease and is expected to be Benign. Please refer to the Clinical Summary section of this report and PMID: 30260545, 30241959 for guidelines on which co-occurring variants may be clinically relevant. ClinVar contains an entry for this variant (Variation ID: 5370). Invitae Evidence Modeling of protein sequence and biophysical properties (such as structural, functional, and spatial information, amino acid conservation, physicochemical variation, residue mobility, and thermodynamic stability) has been performed for this missense variant. However, the output from this modeling did not meet the statistical confidence thresholds required to predict the impact of this variant on NPHS2 protein function. This variant has been shown to alter NPHS2 (podocin) protein function, resulting in podocin mislocalization only when in combination with certain variants, but not in isolation (PMID: 24509478). In summary, this variant is expected to be Benign in homozygosity or in combination with the majority of NPHS2 variants. However, when in trans with certain specific NPHS2 variants, it may be associated with disease. The clinical significance of this variant when in combination with a novel NPHS2 variant is difficult to predict. Therefore, it has been classified as a Variant of Uncertain Significance. -
Inborn genetic diseases Pathogenic:1
The c.686G>A (p.R229Q) alteration is located in exon 5 (coding exon 5) of the NPHS2 gene. This alteration results from a G to A substitution at nucleotide position 686, causing the arginine (R) at amino acid position 229 to be replaced by a glutamine (Q)._x000D_ _x000D_ _x000D_ _x000D_ Based on the available evidence, the NPHS2 c.686G>A (p.R229Q) alteration is classified as likely pathogenic when occurring in trans with certain pathogenic or likely pathogenic NPHS2 alterations in exon 7 or 8. Based on data from gnomAD, the A allele has an overall frequency of 3.025% (8538/282294) total alleles studied. The highest observed frequency was 6.840% (1715/25074) of European (Finnish) alleles. This alteration has been reported in trans with a second NPHS2 alteration located in exon 7 or 8 in multiple individuals with clinical features consistent with NPHS2-related nephrotic syndrome (Tsukaguchi, 2002; Karle, 2002; Machuca, 2009; Lipska, 2013; Mikó, 2018; Rood, 2019). The clinical presentation for these individuals is less severe than typical and may manifest in adulthood (Mikó, 2018; Rood, 2019). In the homozygous state, this alteration is not disease-causing, and it is unlikely to be disease-causing when in trans with a second NPHS2 alteration in exons 1-6 (Mikó, 2018; Rood, 2019). This amino acid position is well conserved in available vertebrate species. Functional analysis demonstrated decreased nephrin binding to the p.R229Q podocin compared to the wild type (Tsukaguchi, 2002). Additional functional studies suggest that this alteration is pathogenic when in trans with certain NPHS2 mutations due to altered heterodimerization and mislocalization of the encoded p.R229Q podocin (Tory, 2014). The in silico prediction for this alteration is inconclusive. Based on the available evidence, this alteration is classified as likely pathogenic. -
NPHS2-related disorder Pathogenic:1
The NPHS2 c.686G>A variant is predicted to result in the amino acid substitution p.Arg229Gln. This variant is reported in 6.8% of alleles in individuals of European (Finnish) descent in gnomAD. It is not pathogenic in the homozygous state but has been associated with late-onset steroid-resistant nephrotic syndrome (SRNS) only when in trans with certain pathogenic NPHS2 variants within or near oligomerization sites (Mikó et al. 2018. PubMed ID: 30260545). Pathogenic oligomerization-site variants (located within residues 283-313 and 332-348; exons 7-8) have been found to exert a deleterious dominant-negative effect on p.Arg229Gln podocin, but behave as recessive alleles when associated with control podocin. Therefore, the pathogenicity of the p.Arg229Gln variant should be carefully interpreted in the context of an individual’s genotype of the NPHS2 gene as well as clinical findings, family history and other laboratory data (Mikó et al. 2018. PubMed ID: 30260545). Taken together, this variant is interpreted as pathogenic. -
not specified Uncertain:1
Variant summary: NPHS2 c.686G>A (p.Arg229Gln) results in a conservative amino acid change located in the Band 7 domain (IPR001107) of the encoded protein sequence. Three of five in-silico tools predict a benign effect of the variant on protein function. The variant allele was found at a frequency of 0.031 in 253464 control chromosomes in the gnomAD database, including 170 homozygotes. The observed variant frequency is approximately 17 fold of the estimated maximal expected allele frequency for a pathogenic variant in NPHS2 causing Nephrotic Syndrome, Type 2 phenotype (0.0018). c.686G>A has been reported in the literature in multiple compound heterozygous individuals affected with Nephrotic Syndrome Type 2 or End-Stage Renal Disease, but also in unaffected individuals (e.g. Zhang_2004, Hinkes_2007, Machuca_2009, Kerti_2013, McCarthy_2013, Benetti_2014, Guaragna_2015, Ottlewski_2019, Baylarov_2020, Riguetti_2020, Internal data). The variant has also been observed in a homozygous state in unaffected individuals (e.g. Kerti_2013, Machuca_2009). Co-occurrences in patients harboring homozygous or compound heterozygous disease causing variants have also been reported, e.g. in the NPHS1 gene (Hinkes_2007), in the PAX2 gene (a de-novo truncating variant; Kerti_2013), and in the PRPRO gene (homozygous splice variant; Ozaltin_2011). An extensive study performed by Tory_2014, concluded that the pathogenicity of Arg229Gln is highly dependent on the NPHS2 variant observed in trans, as missense mutations affecting Ala284, Ala288, Arg291, Ala297, Glu310, Leu327 or Gln328 (located in exons 7 and 8) were enriched in cases carrying Arg229Gln, however, variants located in exons 1-6 in trans with R299Q were less likely to be deleterious. When the Arg229Gln variant was co-expressed with certain variants (i.e. Ala284Val, Ala288Thr, Arg291Trp, Ala297Val or Glu310Lys), it was demonstrated to have a deleterious effect due to altered heterodimerization and mislocalization of the encoded p.Arg229Gln podocin, mimicking a dominant-negative effect, while other variants (e.g. Arg138Gln, Arg238Ser) did not alter the localization of p.Arg229Gln podocin (Tory_2014). In addition, another publication reported experimental evidence on protein function, and found decreased nephrin binding to the R229Q podocin (Tsukaguchi_2002). A review by Miko_2018 concluded that p.R229Q trans-associations can be considered pathogenic if the variant in trans meets the following criteria: 1) it affects residues 270-351 and alters but does not disrupt oligomerization, 2) its p.R229Q association is found in a family with slowly progressing focal segmental glomerulosclerosis, but is expected to be rare in the general population (<1:10 million); they also reported that over 15% of p.R229Q associations identified so far in patients are benign. A recent population-genetic study also confirmed that the variant is subject to interallelic interactions which results in an incomplete penetrance (Miko_2021). The following publications have been ascertained in the context of this evaluation (PMID: 24227627, 26211502, 19520069, 16900088, 19268410, 20798252, 24969201, 19145239, 23349334, 17371932, 21355056, 11805166, 23800802, 18499321, 23645318, 15954915, 16481888, 17942957, 30241959, 18823551, 12464671, 15327385, 26420286, 21415313, 28529802, 33193607, 21722858, 24509478, 32129207, 31738409, 31027891, 34405919, 30260545, 34853150).ClinVar contains an entry for this variant (Variation ID: 5370). Based on the evidence outlined above, the variant was classified as VUS-possibly pathogenic. -
Proteinuria Uncertain:1
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Focal segmental glomerulosclerosis Benign:1
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Nephrotic syndrome, type 2, susceptibility to Other:1
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Nephrotic syndrome Other:1
This individual is heterozygous for a known variant, c.686G>A p.(Arg229Gln), in the NPHS2 gene. The c.686G>A p.(Arg229Gln) variant is reported to be associated with adult-onset steroid-resistant nephrotic syndrome and the development of FSGS when in conjunction with a second pathogenic allele (Tsukaguchi et al 2002 J Clin Invest110: 1659-1666 & Machuca et al 2009 Kidney Int 75: 727-735). It is common amongst Western European populations with an allelic frequency of 3.6% and has variable penetrance. In vitro studies show the NPHS2 p.Arg229Gln variant decreases nephrin binding to podocin and it is considered to play a role in the pathogenesis of SRNS when it is in trans with another pathogenic variant (Machuca et al 2009 Kidney Int 75:727-735). The c.686G>A variant is considered to be a risk factor allele, conferring susceptibility to nephrotic syndrome type 2. -
Computational scores
Source:
Splicing
Find out detailed SpliceAI scores and Pangolin per-transcript scores at