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Hum Mol Genet. 2016 Mar 15;25(6):1152-64. doi: 10.1093/hmg/ddv638. Epub 2016 Jan 5.

Three-layered proteomic characterization of a novel ACTN4 mutation unravels its pathogenic potential in FSGS.

Author information

1
Department II of Internal Medicine and Center for Molecular Medicine Cologne, University of Cologne, Cologne, Germany.
2
Department II of Internal Medicine and Center for Molecular Medicine Cologne, University of Cologne, Cologne, Germany, Department of Pediatrics.
3
Department II of Internal Medicine and Center for Molecular Medicine Cologne, University of Cologne, Cologne, Germany, Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases (CECAD) and Systems Biology of Ageing Cologne, University of Cologne, Cologne, Germany.
4
Department of Pediatrics.
5
Cologne Center for Genomics (CCG).
6
Institute of Human Genetics, Cologne Center for Genomics (CCG).
7
Department of Radiology.
8
Institute of Human Genetics.
9
Institute for Genetics, Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases (CECAD) and.
10
Department II of Internal Medicine and Center for Molecular Medicine Cologne, University of Cologne, Cologne, Germany, Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases (CECAD) and Systems Biology of Ageing Cologne, University of Cologne, Cologne, Germany markus.rinschen@uk-koeln.de bodo.beck@uk-koeln.de.
11
Institute of Human Genetics, markus.rinschen@uk-koeln.de bodo.beck@uk-koeln.de.

Abstract

Genetic diseases constitute the most important cause for end-stage renal disease in children and adolescents. Mutations in the ACTN4 gene, encoding the actin-binding protein α-actinin-4, are a rare cause of autosomal dominant familial focal segmental glomerulosclerosis (FSGS). Here, we report the identification of a novel, disease-causing ACTN4 mutation (p.G195D, de novo) in a sporadic case of childhood FSGS using next generation sequencing. Proteome analysis by quantitative mass spectrometry (MS) of patient-derived urinary epithelial cells indicated that ACTN4 levels were significantly decreased when compared with healthy controls. By resolving the peptide bearing the mutated residue, we could proof that the mutant protein is less abundant when compared with the wild-type protein. Further analyses revealed that the decreased stability of p.G195D is associated with increased ubiquitylation in the vicinity of the mutation site. We next defined the ACTN4 interactome, which was predominantly composed of cytoskeletal modulators and LIM domain-containing proteins. Interestingly, this entire group of proteins, including several highly specific ACTN4 interactors, was globally decreased in the patient-derived cells. Taken together, these data suggest a mechanistic link between ACTN4 instability and proteome perturbations of the ACTN4 interactome. Our findings advance the understanding of dominant effects exerted by ACTN4 mutations in FSGS. This study illustrates the potential of genomics and complementary, high-resolution proteomics analyses to study the pathogenicity of rare gene variants.

PMID:
26740551
DOI:
10.1093/hmg/ddv638
[Indexed for MEDLINE]

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