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NPHS2 STOMATIN FAMILY MEMBER, PODOCIN; NPHS2

NPHS2 STOMATIN FAMILY MEMBER, PODOCIN; NPHS2

Alternative titles; symbolsPODOCIN; PDCNHGNC Approved Gene Symbol: NPHS2Cytogenetic location: 1q25.2 Genomic coordinates (GRCh38): 1:179,550,538-179,575,986 ...

Alternative titles; symbols

  • PODOCIN; PDCN

HGNC Approved Gene Symbol: NPHS2

Cytogenetic location: 1q25.2 Genomic coordinates (GRCh38): 1:179,550,538-179,575,986 (from NCBI)

▼ Description
The NPHS2 gene encodes podocin, a protein that is expressed at the insertion of the slit diaphragm in the renal glomerulus. Podocytes are specialized epithelial cells covering the basement membrane of the renal glomerulus, and they mediate glomerular filtration through the slit diaphragm. Podocin is thus crucial in the establishment of the glomerular filtration barrier (Roselli et al., 2004).

▼ Cloning and Expression
Boute et al. (2000) used positional cloning to identify the causative gene in a form of steroid-resistant nephrotic syndrome (NPHS2; 600995) mapping to chromosome 1q25-q31. The NPHS2 gene encodes an integral membrane protein that they named podocin. Boute et al. (2000) isolated an NPHS2 cDNA from a human fetal kidney cDNA library. The full-length NPHS2 cDNA encodes a putative 383-amino acid protein of approximately 42 kD. The 3-prime untranslated region contains an atypical polyadenylation signal situated upstream of the poly(A) tail. One of 2 overlapping consensus polyadenylation signals located downstream of the aforementioned signal can also be used. Database comparisons showed a region of extensive similarity between the central region of podocin and proteins of the band-7-stomatin (see 133090) family. The strongest homology was found with human stomatin (47% identity over 253 amino acids) and Caenorhabditis elegans MEC-2 (44% identity over 275 amino acids). Analysis of the amino acid sequence suggested that NPHS2 encodes an integral membrane protein with 1 transmembrane domain and a C-terminal cytoplasmic tail. The transmembrane domain and most of the cytoplasmic tail are homologous to the corresponding regions of the stomatin family proteins. Furthermore, the cysteine residue located 4 amino acids upstream of the transmembrane domain, which is conserved among stomatins and MEC-2 and has been shown to be the major palmitoylation site in stomatin, was also conserved in podocin. The N and C termini of podocin showed no homology with any known protein sequence. Northern blot analysis detected a 2-kb transcript strongly expressed in human fetal and adult kidney, with no signal being observed in other tissues. Boute et al. (2000) performed in situ hybridization on normal fetal and postnatal kidney samples and observed strong signals exclusively in the glomeruli. In the fetuses no signal was detected in the earlier stages of nephron development, whereas intense signals were seen in the future podocytes of the inferior segment of the S-body, concomitant with vascularization of the inferior cleft. Under high magnification, transcript expression was restricted to within the podocytes, at the periphery of the glomerular tuft.

Schurek et al. (2014) presented evidence that podocin has a hairpin-like topology, with both N- and C-terminal domains facing the cytoplasm. A stretch of approximately 20 hydrophobic amino acids dips into the inner leaflet of the plasma membrane. The kink of the hairpin, provided by highly conserved pro118, ensures membrane proximity of the C-terminal PHB domain. Palmitoylation of 2 conserved cysteine residues fixes membrane proximity of the PHB domain.

▼ Mapping
The NPHS2 gene maps to chromosome 1q25-q31 (Boute et al., 2000).

Stumpf (2022) mapped the NPHS2 gene to chromosome 1q25.2 based on an alignment of the NPHS2 sequence (GenBank BC029141) with the genomic sequence (GRCh38).

▼ Gene Function
Hereditary nephrotic syndrome is a heterogeneous disease, characterized by heavy proteinuria and renal failure. Mutations of the genes encoding for nephrin (NPHS1; 602716) and podocin lead to early onset of heavy proteinuria and rapid progression to end-stage renal disease. Huber et al. (2003) showed that wildtype podocin is targeted to the plasma membrane and forms homooligomers. The association of podocin with specialized lipid raft microdomains of the plasma membrane was a prerequisite for recruitment of nephrin into rafts. In contrast, the R138Q (604766.0001) and R138X (604766.0002) mutations blocked recruitment of nephrin into rafts, because the R138Q mutant was retained in the endoplasmic reticulum, and the R138X mutant failed to associate with rafts despite their presence in the plasma membrane. Neither mutant augmented nephrin signaling, suggesting that lipid raft targeting may facilitate nephrin signaling.

Using wildtype and mutant (pro118 to leu, P118L) human podocin or equivalent mouse and C. elegans mutant proteins, Schurek et al. (2014) found that P118L-substituted podocin was predominantly retained in the endoplasmic reticulum. The portion of mutant podocin that was transported to the plasma membrane had an extracellular C terminus that was subject to glycosylation. Wildtype, but not P118L mutant, podocin associated with membrane rafts and bound cholesterol. When expressed in Xenopus oocytes, wildtype but not mutant mouse podocin associated with the human ion channel TRPC6 (603652) and augmented TRPC6 currents. Schurek et al. (2014) concluded that the P118L mutation changes the hairpin-like domain of podocin to a transmembrane domain, significantly changing the interaction of podocin with lipid rafts and channels involved in kidney filtration barrier function.

▼ Molecular Genetics
In 14 families with autosomal recessive steroid-resistant nephrotic syndrome (NPHS2; 600995), Boute et al. (2000) identified 10 different mutations in the NPHS2 gene (604766.0001-604766.0010). Three mutations resulted in a frameshift or premature stop codon. Six were missense mutations; R138Q (604766.0001) was identified in 6 different families. The findings demonstrated a crucial role for podocin in the function of the glomerular filtration barrier.

Tsukaguchi et al. (2002) identified a polymorphism in the NPHS2 gene (R229Q; 604766.0011) that contributed to the development of late-onset renal disease characterized by proteinuria and focal segmental glomerulosclerosis (FSGS) in 6 families. In vitro functional expression assays indicated that the R229Q mutant protein had decreased binding to nephrin (NPHS1; 602716) (41.5% compared to wildtype), suggesting mild functional impairment. Tsukaguchi et al. (2002) concluded that the R229Q variant itself may not cause disease, but appeared to increase susceptibility to renal disease when combined with another pathogenic NPHS2 mutation.

▼ Genotype/Phenotype Correlations
In 7 (18%) of 38 Hungarian patients with childhood-onset nephrotic-range proteinuria, Kerti et al. (2013) identified biallelic mutations in the NPHS2 gene. Three patients had the R138Q mutation on 1 allele and a truncating mutation on the second allele, 1 patient was homozygous for the R138Q mutation, and 1 patient had 2 truncating mutations. All 5 of these patients had the classic phenotype, with onset of the disorder in early childhood and progression to end-stage renal disease in 4 patients by age 10 years. Two patients with a much milder phenotype with only proteinuria and normal renal function at ages 18 and 31 years, respectively, carried a V290M substitution (604766.0012). One was homozygous for V290M and the other was compound heterozygous for V290M and R138Q. A third patient with more classic severe NPHS2 resulting in end-stage renal disease at age 8 years carried the V290M variant in the heterozygous state; a second mutation was presumably present but not detected. Kerti et al. (2013) suggested that screening for the V290M mutation should be included for patients with late-onset nephrotic syndrome.

Tory et al. (2014) presented evidence that the R229Q variant (604766.0011) is pathogenic only when combined with a particular type of NPHS2 mutation on the other allele. Among 129 unaffected parents of patients with nephrotic syndrome-2, 6 (4.7%) were found to be compound heterozygous for R229Q on 1 allele and R138Q (604766.0001), R138X (604776.0002), R168H, c.534+1G-T, or R238S on the other allele. These findings clearly demonstrated incomplete penetrance of nephrotic syndrome in individuals with a certain genotype R229Q/mut, when the second mutation occurs in exons 1 to 6 of the NPHS2 gene. In a retrospective analysis of 318 families with NPHS2, there were significantly more patients who were compound heterozygous for R229Q and a mutation affecting exons 7 and 8 of the NPHS2 gene (see, e.g., R291W; 604766.0010) compared to patients who were compound heterozygous for R229Q and a mutation in exons 1 to 6 of the NPHS2 gene (p = 1.2 x 10(-35)). Missense mutations affecting residues ala284, ala288, arg291, ala297, glu310, leu327, or gln328 were enriched in cases with the R229Q mutation. All of these associated mutations in exons 7 and 8 were 3-prime terminal mutations. Transfection studies in human podocyte cell lines showed that the R229Q variant protein localized properly to the plasma membrane when coexpressed with wildtype or variants affecting exons 1 to 6 (R238S or V290M). However, the R229Q protein was retained in the cytoplasm when coexpressed with mutations affecting exons 7 and 8. These interactions mimicked a dominant-negative effect of these mutations on the R229Q variant protein. Structural modeling showed that the combination of R229Q and the pathogenic variants leads to an altered mode of dimerization that likely contributes to the retention of R229Q podocin within cytoplasmic compartments. Tory et al. (2014) noted that these findings have implications for genetic counseling, since the transmission pattern of the disorder may be mutation-dependent.

▼ Animal Model
Roselli et al. (2004) generated podocin-deficient mice to investigate the role of podocin in renal function and in the development of SRN1 and glomerulosclerosis. Podocin-null mice developed proteinuria during the antenatal period and died a few days after birth from renal failure caused by massive mesangial sclerosis. Electron microscopy revealed the extensive fusion of podocyte foot processes and the lack of a slit diaphragm in the remaining foot process junctions. Using real-time PCR and immunolabeling, Roselli et al. (2004) found that expression of other slit diaphragm components was modified in podocin-null kidneys: nephrin was downregulated, whereas ZO1 (601009) and CD2AP (604241) appeared to be upregulated. The progression of renal disease, as well as the presence or absence of renal vascular lesions, depended on the genetic background.

▼ ALLELIC VARIANTS ( 12 Selected Examples):

.0001 NEPHROTIC SYNDROME, TYPE 2
NPHS2, ARG138GLN
In 6 families with steroid-resistant nephrotic syndrome type 2 (NPHS2; 600995), 3 of which were previously reported by Fuchshuber et al. (1995), Boute et al. (2000) identified a 413G-A transition in exon 3 of the NPHS2 gene, resulting in an arg138-to-gln (R138Q) substitution. The mutation was found in homozygosity in 4 families and in compound heterozygosity in another (see 604766.0009); in 1 family, only a paternal R138Q mutation was detected. This mutation causes the replacement of a residue that is highly conserved among stomatin-like protein family members and is probably crucial for podocin function.

Caridi et al. (2001) found a homozygous R138Q mutation in 3 patients with sporadic steroid-resistant nephrotic syndrome with focal segmental glomerulosclerosis. Onset occurred before age 2 years and all progressed to renal failure necessitating transplantation.

Koziell et al. (2002) identified a homozygous R138Q mutation in an English patient with a severe form of NPHS2 with congenital onset.

.0002 NEPHROTIC SYNDROME, TYPE 2
NPHS2, ARG138TER
In a consanguineous family with autosomal recessive steroid-resistant nephrotic syndrome (600995) previously reported by Fuchshuber et al. (1995), Boute et al. (2000) identified a homozygous 412C-T transition in exon 3 of the NPHS2 gene, resulting in an arg138-to-ter (R138X) substitution.

Frishberg et al. (2002) identified a homozygous R138X mutation in 15 (55%) of 27 children with steroid-responsive nephrotic syndrome. All were of Arab-Israeli descent, and haplotype analysis indicated a founder effect. Renal histology included diffuse mesangial proliferation and focal segmental glomerulosclerosis.

Becker-Cohen et al. (2007) reported a 9-year-old Arab girl with NPHS2 due to a homozygous R138X mutation. She presented with recurrent nephrotic syndrome 4 years after renal transplantation from a deceased donor and was treated with plasmapheresis with a partial response. However, renal histology did not demonstrate glomerular immunoglobulin deposition, and an extensive search for anti-podocin antibodies based on indirect Western blot was negative. The findings confirmed the possibility of post transplantation nephrotic syndrome in patients with NPHS2 mutations, but the lack of immunoglobulin deposition and anti-podocin antibodies implicated another pathogenic mechanism for disease recurrence.

.0003 NEPHROTIC SYNDROME, TYPE 2
NPHS2, 1-BP INS, 104G
In a family with autosomal recessive steroid-resistant nephrotic syndrome (600995), Boute et al. (2000) identified a frameshift mutation in exon 1 of the NPHS2 gene, which resulted from the insertion of a G at nucleotide 104 (104insG). This mutation was found in compound heterozygosity with a second frameshift mutation (see 604766.0004).

.0004 NEPHROTIC SYNDROME, TYPE 2
NPHS2, 1-BP DEL, 419G
In a family with steroid-resistant nephrotic syndrome (600995), Boute et al. (2000) identified a frameshift mutation, a deletion of a G at nucleotide 419 (419delG) in exon 3 of the NPHS2 gene. This mutation was found in compound heterozygosity with another frameshift mutation (see 604766.0003).

Caridi et al. (2001) found a homozygous 419delG mutation in 3 Italian patients with sporadic steroid-resistant nephrotic syndrome with focal segmental glomerulosclerosis. Onset occurred before age 4 years and all progressed to renal failure necessitating transplantation. Haplotype analysis indicated a founder effect.

.0005 NEPHROTIC SYNDROME, TYPE 2
NPHS2, 2-BP DEL, 855AA
In a family with autosomal recessive steroid-resistant nephrotic syndrome (600995) previously reported by Fuchshuber et al. (1995), Boute et al. (2000) identified a frameshift mutation on the paternal allele of the NPHS2 gene of an affected patient. The frameshift resulted from deletion of 2 adenines at nucleotide 855 and 856 in exon 7. No mutation was found on the other allele.

.0006 NEPHROTIC SYNDROME, TYPE 2
NPHS2, PRO20LEU
In a consanguineous family with autosomal recessive steroid-resistant nephrotic syndrome (600995), Boute et al. (2000) identified a homozygous 59C-T transition in exon 1 of the NPHS2 gene, resulting in a pro20-to-leu (P20L) substitution.

Caridi et al. (2003) identified a heterozygous P20L mutation in 5 unrelated patients with a slightly milder form of NPHS2, 3 of whom showed response to steroid treatment. Although the authors could not exclude another pathogenic mutation, they suggested that heterozygous NPHS2 mutations may be associated with a milder phenotype.

.0007 NEPHROTIC SYNDROME, TYPE 2
NPHS2, GLY92CYS
In a family with autosomal recessive steroid-resistant nephrotic syndrome (600995) previously reported by Fuchshuber et al. (1995), Boute et al. (2000) identified a 274G-T transversion in exon 1 of the NPHS2 gene resulting in a gly92-to-cys (G92C) substitution. This mutation was found on the paternal allele; no mutation was identified on the maternal allele. In addition to being a missense mutation, this mutation also alters the last nucleotide of exon 1 and thus probably alters splicing.

.0008 NEPHROTIC SYNDROME, TYPE 2
NPHS2, ASP160GLY
In a consanguineous family with autosomal recessive steroid-resistant nephrotic syndrome (600995), Boute et al. (2000) identified a homozygous 479A-G transition in exon 4 of the NPHS2 gene, which resulted in an asp160-to-gly (D160G) substitution. This mutation causes the replacement of a residue that is highly conserved among stomatin-like protein family members and that is probably crucial for podocin function.

.0009 NEPHROTIC SYNDROME, TYPE 2
NPHS2, VAL180MET
In a consanguineous family with steroid-resistant nephrotic syndrome (600995) previously reported by Fuchshuber et al. (1996), Boute et al. (2000) identified a homozygous 538G-A transition in exon 5 of the NPHS2 gene, which resulted in a val180-to-met (V180M) substitution. In another family, it was identified in compound heterozygosity in conjunction with R138Q (604766.0001).

.0010 NEPHROTIC SYNDROME, TYPE 2
NPHS2, ARG291TRP
In a family with autosomal recessive steroid-resistant nephrotic syndrome (600995) previously reported by Fuchshuber et al. (1995), Boute et al. (2000) identified an 871C-T transition in exon 7 of the NPHS2 gene, which resulted in an arg291-to-trp (R291W) substitution. This mutation was identified on the paternal allele; a mutation on the maternal allele was not identified.

.0011 NEPHROTIC SYNDROME, TYPE 2, SUSCEPTIBILITY TO
NPHS2, ARG229GLN (rs61747728)
In a large Brazilian family with adolescent or adult onset of renal disease characterized by proteinuria leading to end-stage renal failure in some and showing focal segmental glomerulosclerosis on biopsy (NPHS2; 600995), Tsukaguchi et al. (2002) identified compound heterozygosity for 2 variants in the NPHS2 gene: a 755G-A transition in exon 5 of the NPHS2 gene resulting in an arg229-to-gln (R229Q) substitution, and R291W (604766.0010). The age at disease onset ranged from 18 to 36 years, and 2 patients had end-stage renal disease in their thirties. The R229Q allele was identified in 5 additional families with a similarly mild disorder. All affected individuals were compound heterozygous for R229Q and another putative pathogenic NPHS2 mutation (see, e.g., R138Q; 604766.0001). The R229Q variant was found in 3.6% of controls, indicating that it is a polymorphism. Haplotype analysis of patients and controls carrying the variant indicated a founder effect. However, in vitro functional expression assays indicated that the R229Q mutant protein had decreased binding to nephrin (NPHS1; 602716) (41.5% compared to wildtype), suggesting mild functional impairment. Tsukaguchi et al. (2002) concluded that the R229Q variant itself may not cause disease, but may increase susceptibility to renal disease when combined with another pathogenic NPHS2 mutation.

Ozaltin et al. (2011) reported a Turkish girl, born of consanguineous parents, who had NPHS6 (614196) due to a homozygous splice site mutation in the PTPRO gene (600579.0002). She also carried a heterozygous R229Q mutation in the podocin gene. She had onset of steroid-resistant nephrotic syndrome at age 14, which progressed to end-stage renal disease necessitating renal transplant at age 18. Her phenotype was more severe compared to her 2 sibs who carried the homozygous PTPRO mutation but not the R229Q mutation. Ozaltin et al. (2011) suggested that the R229Q mutation may have exacerbated the clinical picture in this patient.

Kerti et al. (2013) reported a 37-year-old Hungarian man with onset of subnephrotic proteinuria (see NPHS2, 600995) at age 7 months. He later developed hypertension and had persistent steroid-resistant nephrotic-range proteinuria. Renal biopsy at age 20 years showed FSGS, and he developed end-stage renal disease necessitating renal transplant at age 33. Molecular studies showed that he was homozygous for the R229Q variant. However, his unaffected father and brother were also homozygous for R229Q. The allele frequency of R229Q was found to be 3% among 212 Hungarian controls. The proband also had atrophy of the optic nerve head and a morning glory disc anomaly associated with nystagmus and poor vision. He was found to carry a de novo truncating mutation in the PAX2 gene (167409.0002), which is mutated in autosomal dominant papillorenal syndrome (120330). Kerti et al. (2013) concluded that homozygosity for R229Q does not cause FSGS in itself, but may have a modifier effect when present with another pathogenic mutation.

Tory et al. (2014) presented evidence that the R229Q variant is pathogenic only when combined with a particular type of NPHS2 mutation on the other allele. Transfection studies in human podocyte cell lines showed that the R229Q variant protein localized properly to the plasma membrane when coexpressed with wildtype or variants affecting exons 1 to 6 (R238S or V290M). However, the R229Q protein was retained in the cytoplasm when coexpressed with mutations affecting exons 7 and 8. These interactions mimicked a dominant-negative effect of these mutations on the R229Q variant protein. Structural modeling showed that the combination of R229Q and the pathogenic variants leads to an altered mode of dimerization that likely contributes to the retention of R229Q podocin within cytoplasmic compartments. Tory et al. (2014) noted that these findings have implications for genetic counseling, since the transmission pattern of the disorder may be mutation-dependent.

.0012 NEPHROTIC SYNDROME, TYPE 2, SUSCEPTIBILITY TO
NPHS2, VAL290MET
In 2 individuals with nephrotic-range proteinuria (NPHS2; 600995) detected in a school screening program Kerti et al. (2013) identified a c.868G-A transition in exon 7 of the NPHS2 gene, resulting in a val290-to-met (V290M) substitution. One individual was homozygous for the variant and the other was compound heterozygous for V290M and R138Q (604766.0001). Both individuals had normal renal function at ages 18 and 31 years, respectively. Renal biopsy showed FSGS in the homozygous individual and minimal disease change in the compound heterozygous individual. A third patient with more classic severe NPHS2 resulting in end-stage renal disease at age 8 years carried the V290M variant in the heterozygous state; a second mutation was presumably present but not detected. The V290M variant was not found in 83 French patients with late-onset nephrotic syndrome. However, 2 (1.1%) of 95 European patients with late-onset nephrotic syndrome carried V290M: a German individual carried V290M in compound heterozygosity with a splice site mutation in the NPHS2 gene, and a Turkish patient was heterozygous for V290M. Haplotype analysis suggested a founder effect for V290M. Functional studies of the variant were not performed. Kerti et al. (2013) suggested that screening for the V290M mutation should be included for patients with late-onset nephrotic syndrome.

Tags: 1q25.2