Single-Molecule Real-Time Sequencing for MUC1 VNTR Variation to Improve Autosomal Dominant Tubulointerstitial Kidney Disease Diagnosis

BACKGROUND: ADTKD-MUC1 is caused by frameshift mutations in the MUC1 gene, producing a frameshifted neoprotein (MUC1fs) toxic to kidney cells. The gene's variable number of tandem repeats (VNTR), with ∼80% guanine/cytosine content, has made it largely inaccessible to standard short-read sequencing, leaving the reference sequence and natural variation poorly defined and complicating mutation detection. METHODS: Using Single Molecule, Real-Time (SMRT) sequencing, we characterized MUC1 VNTR in 300 individuals, including 279 from 143 families suspected of having ADTKD-MUC1, assessing VNTR length, repeat structure, and frameshift mutations. Results were compared with the CLIA-approved probe-extension assay, detecting the prevalent 59dupC mutation, and with MUC1fs immunohistochemistry, which detects the pathogenic protein independent of the underlying genomic change. RESULTS: We identified 215 unique VNTR alleles composed of 80 distinct repeat units, 46 (58%) of which were novel, and nine distinct frameshift mutations present on 52 mutated alleles. Overall, MUC1 frameshift mutations were identified in 71 of 143 families (50%) with suspected ADTKD-MUC1, comprising 135 affected individuals (48%). The SMRT assay outperformed the probe-extension assay by identifying frameshift mutations in two families with previously inconclusive results and in eight additional families whose mutations were undetectable by the probe-extension design. When successful, SMRT assay showed 100% concordance with probe-extension assay at the family level and 98% at the individual level, with discordance attributable to allelic dropout inherent to long-read sequencing. Analysis of the mutational spectrum confirmed 59dupC as the most prevalent mutation, affecting ∼90% of families, while the other eight mutation types occurred at most twice. CONCLUSIONS: The SMRT assay outperformed the CLIA-approved probe-extension assay by detecting essentially all VNTR-associated frameshift mutations. The probe-extension assay identified ∼90% of affected families. MUC1fs immunohistochemistry added diagnostic value in genetically unresolved cases by detecting the pathogenic protein independent of the underlying mutation.

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Journal
Journal of the American Society of Nephrology
Published
2026-04-10
DOI
https://doi.org/10.1681/asn.0000001103
Primary Topic
Renal Diseases and Glomerulopathies
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article
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article

Single-Molecule Real-Time Sequencing for MUC1 VNTR Variation to Improve Autosomal Dominant Tubulointerstitial Kidney Disease Diagnosis

Jana Sovová, Omri Teltsh, C. C. Deltas, Martina Živná et al.
Journal of the American Society of Nephrology
Renal Diseases and Glomerulopathies
article

Single-Molecule Real-Time Sequencing for MUC1 VNTR Variation to Improve Autosomal Dominant Tubulointerstitial Kidney Disease Diagnosis

Jana Sovová, Omri Teltsh, C. C. Deltas, Martina Živná, Hana Hartmannová, Alena Vrbacká, Viktor Stránecký, Brendan Blumenstiel, Silvie Rajnochová-Bloudíčková, Lenka Mrázová, Romana Ryšavá, Kendrah Kidd, Anna Přistoupilová, Petr Vyleťal, Tereza Kmochová, Christoforos Stavrou, Václav Janoušek, Helena Trešlová, Gregory Papagregoriou, Gianpiero L. Cavalleri, Kateřina Hodaňová, Martin Radina, Dita Mušálková, Anthony J. Bleyer, Peter J. Conlon, Diana M. Toledo, Michelle Clince, Matthew DeFelice, Stanislav Kmoch, Veronika Barešová, O. Viklicky, Ibrahim Bitar, Klára Svojšová, Heidi A. Bleyer, Antonio Güemes Sánchez, Lauren Martin, Sofía Jorge, Colm Rowan, Innet Lajtmanov, J. Revez Lopes, Marina Distefano, Elhussein Elhassan, Abby Taylor
article en

Abstract

BACKGROUND: ADTKD-MUC1 is caused by frameshift mutations in the MUC1 gene, producing a frameshifted neoprotein (MUC1fs) toxic to kidney cells. The gene's variable number of tandem repeats (VNTR), with ∼80% guanine/cytosine content, has made it largely inaccessible to standard short-read sequencing, leaving the reference sequence and natural variation poorly defined and complicating mutation detection. METHODS: Using Single Molecule, Real-Time (SMRT) sequencing, we characterized MUC1 VNTR in 300 individuals, including 279 from 143 families suspected of having ADTKD-MUC1, assessing VNTR length, repeat structure, and frameshift mutations. Results were compared with the CLIA-approved probe-extension assay, detecting the prevalent 59dupC mutation, and with MUC1fs immunohistochemistry, which detects the pathogenic protein independent of the underlying genomic change. RESULTS: We identified 215 unique VNTR alleles composed of 80 distinct repeat units, 46 (58%) of which were novel, and nine distinct frameshift mutations present on 52 mutated alleles. Overall, MUC1 frameshift mutations were identified in 71 of 143 families (50%) with suspected ADTKD-MUC1, comprising 135 affected individuals (48%). The SMRT assay outperformed the probe-extension assay by identifying frameshift mutations in two families with previously inconclusive results and in eight additional families whose mutations were undetectable by the probe-extension design. When successful, SMRT assay showed 100% concordance with probe-extension assay at the family level and 98% at the individual level, with discordance attributable to allelic dropout inherent to long-read sequencing. Analysis of the mutational spectrum confirmed 59dupC as the most prevalent mutation, affecting ∼90% of families, while the other eight mutation types occurred at most twice. CONCLUSIONS: The SMRT assay outperformed the CLIA-approved probe-extension assay by detecting essentially all VNTR-associated frameshift mutations. The probe-extension assay identified ∼90% of affected families. MUC1fs immunohistochemistry added diagnostic value in genetically unresolved cases by detecting the pathogenic protein independent of the underlying mutation.

Journal of the American Society of Nephrology
Broad Institute (US), Royal College of Surgeons in Ireland (IE), Charles University (CZ), Beaumont Hospital (IE), University of Cyprus (CY), Institute of Clinical and Experimental Medicine (CZ), B. Braun (Czechia) (CZ), Hospital de Santa Maria (PT), General University Hospital in Prague (CZ), Wake Forest University (US)
Openalex Percentile: Top 21%
Renal Diseases and Glomerulopathies
DOISourceOpenAlex
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