title: Co-Inheritance of Functional Podocin Variants with Heterozygous Collagen IV Mutations Predisposes to Renal Failure.
abstract: OBJECTIVE
  A subset of patients who present with proteinuria and are diagnosed with focal segmental glomerulosclerosis (FSGS) have inherited heterozygous COL4A3/A4 mutations and are also diagnosed with thin basement membrane nephropathy (TBMN-OMIM: 141200). Two studies showed that co-inheritance of NPHS2-p.Arg229Gln, a podocin variant, may increase the risk for proteinuria and renal function decline.
  
  
  METHODS
  We hypothesized that additional podocin variants may exert a similar effect. We studied genetically a well-characterized Cypriot TBMN patient cohort by re-sequencing the NPHS2 coding region. We also performed functional studies in cell culture experiments, investigating the interaction of podocin variants with itself and with nephrin.
  
  
  RESULTS
  Potentially disease-modifying podocin variants were searched for by analyzing NPHS2 in 35 'severe' TBMN patients. One non-synonymous variant, p.Glu237Gln, was detected. Both variants, p.Arg229Gln and p.Glu237Gln, were tested in a larger cohort of 122 TBMN patients, who were categorized as 'mild' or 'severe' based on the presence of microscopic hematuria alone or combined with chronic renal failure and/or proteinuria. Seven 'severe' patients carried either of the 2 variants; none was present in the 'mild' patients (p = 0.05, Pearson χ(2)). The 7 carriers belong in 2 families segregating mutation COL4A3-p.Gly1334Glu. Inheritance of the wild-type (WT) and mutant alleles correlated with the phenotype (combined concordance probability 0.003). Immunofluorescence (IF) experiments after dual co-transfection of WT and mutant podocin suggested altered co-localization of mutant homodimers. IF experiments after co-transfection of WT podocin and nephrin showed normal membrane localization, while both podocin variants interfered with normal trafficking, demonstrating perinuclear staining. Immunoprecipitation experiments showed stronger binding of mutant podocin to WT podocin or nephrin.
  
  
  CONCLUSIONS
  The results support the hypothesis that certain hypomorphic podocin variants may act as adverse genetic modifiers when co-inherited with COL4A3/A4 mutations, thus predisposing to FSGS and severe kidney function decline.
authors.affiliation: Molecular Medicine Research Center and Laboratory of Molecular and Medical Genetics, Department of Biological Sciences, University of Cyprus, Nicosia, Cyprus.
authors.forename: Charalambos
authors.initials: C
authors.lastname: Stefanou
authors.forename: Myrtani
authors.initials: M
authors.lastname: Pieri
authors.forename: Isavella
authors.initials: I
authors.lastname: Savva
authors.forename: Georgia
authors.initials: G
authors.lastname: Georgiou
authors.forename: Alkis
authors.initials: A
authors.lastname: Pierides
authors.forename: Konstantinos
authors.initials: K
authors.lastname: Voskarides
authors.forename: Constantinos
authors.initials: C
authors.lastname: Deltas
mesh_terms: D000368:Aged; D000483:Alleles; D001324:Autoantigens / Q000235:genetics*; D015331:Cohort Studies; D024141:Collagen Type IV / Q000235:genetics*; D018450:Disease Progression; D005260:Female; D050533:Glomerular Basement Membrane / Q000473:pathology; D005923:Glomerulosclerosis, Focal Segmental / Q000235:genetics / Q000473:pathology; D006579:Heterozygote; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics*; D007676:Kidney Failure, Chronic / Q000453:epidemiology / Q000235:genetics* / Q000473:pathology; D008297:Male; D008565:Membrane Proteins / Q000235:genetics*; D008875:Middle Aged; D009154:Mutation / Q000235:genetics; D010375:Pedigree; D011507:Proteinuria / Q000453:epidemiology / Q000235:genetics; D012737:Sex Factors
chemical_list: D001324:Autoantigens; D024141:Collagen Type IV; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; C407223:NPHS2 protein; C057112:type IV collagen alpha3 chain