Abstract
ABSTRACTRenal calcification (RCALC) resulting in nephrolithiasis and nephrocalcinosis, which affects ∼10% of adults by 70 years of age, involves environmental and genetic etiologies. Thus, nephrolithiasis and nephrocalcinosis occurs as an inherited disorder in ∼65% of patients, and may be associated with endocrine and metabolic disorders including: primary hyperparathyroidism, hypercalciuria, renal tubular acidosis, cystinuria, and hyperoxaluria. Investigations of families with nephrolithiasis and nephrocalcinosis have identified some causative genes, but further progress is limited as large families are unavailable for genetic studies. We therefore embarked on establishing mouse models for hereditary nephrolithiasis and nephrocalcinosis by performing abdominal X‐rays to identify renal opacities in N‐ethyl‐N‐nitrosourea (ENU)‐mutagenized mice. This identified a mouse with RCALC inherited as an autosomal dominant trait, designated RCALC type 2 (RCALC2). Genomewide mapping located the Rcalc2 locus to a ∼16‐Mbp region on chromosome 11D‐E2 and whole‐exome sequence analysis identified a heterozygous mutation in the DNA polymerase gamma‐2, accessory subunit (Polg2) resulting in a nonsense mutation, Tyr265Stop (Y265X), which co‐segregated with RCALC2. Kidneys of mutant mice (Polg2+ / Y265X) had lower POLG2 mRNA and protein expression, compared to wild‐type littermates (Polg2+/+). The Polg2+/Y265X and Polg2+ / + mice had similar plasma concentrations of sodium, potassium, calcium, phosphate, chloride, urea, creatinine, glucose, and alkaline phosphatase activity; and similar urinary fractional excretion of calcium, phosphate, oxalate, and protein. Polg2 encodes the minor subunit of the mitochondrial DNA (mtDNA) polymerase and the mtDNA content in Polg2+ / Y265X kidneys was reduced compared to Polg2+/+ mice, and cDNA expression profiling revealed differential expression of 26 genes involved in several biological processes including mitochondrial DNA function, apoptosis, and ubiquitination, the complement pathway, and inflammatory pathways. In addition, plasma of Polg2+ / Y265X mice, compared to Polg2+ / + littermates had higher levels of reactive oxygen species. Thus, our studies have identified a mutant mouse model for inherited renal calcification associated with a Polg2 nonsense mutation. © 2018 The Authors. Journal of Bone and Mineral Research Published by Wiley Periodicals, Inc.
Highlights
Renal calcification, which comprises nephrocalcinosis and nephrolithiasis, has a multifactorial etiology involving environmental and genetic determinants.[1]
Studies of families with rare monogenic disorders associated with hypercalciuric nephrolithiasis and/or nephrocalcinosis, such as Bartter syndrome, Dent disease, autosomal dominant hypocalcaemia, and distal renal tubular acidosis have identified mutations in >30 genes involved in calcium transport regulation (Supporting Table 1).(3,12,13) genomewide association studies and targeted sequencing of genes with known roles in calcium and vitamin D metabolism have identified associations between nephrolithiasis and common sequence variants in >10 additional genes (Supporting Table 1).(1,14–16) these account for only $15% to 20% of cases,(3,12) and the identification of further monogenic causes of nephrolithiasis are limited by the unavailability of large families
Our studies have demonstrated that renal calcification in a mutant mouse model is due to a heterozygous germline Tyr265Stop mutation of Polg2, which encodes the POLG2 protein that forms part of a heterotrimeric complex composed of two POLG2 subunits and one catalytic polymerase subunit gamma (POLG) subunit.[39]. Mutations in both POLG and POLG2 cause diseases in humans, 504 GORVIN ET AL
Summary
Renal calcification, which comprises nephrocalcinosis and nephrolithiasis (kidney stones), has a multifactorial etiology involving environmental and genetic determinants.[1]. Studies of families with rare monogenic disorders associated with hypercalciuric nephrolithiasis and/or nephrocalcinosis, such as Bartter syndrome, Dent disease, autosomal dominant hypocalcaemia, and distal renal tubular acidosis have identified mutations in >30 genes involved in calcium transport regulation (Supporting Table 1).(3,12,13) genomewide association studies and targeted sequencing of genes with known roles in calcium and vitamin D metabolism have identified associations between nephrolithiasis and common sequence variants in >10 additional genes (Supporting Table 1).(1,14–16) these account for only $15% to 20% of cases,(3,12) and the identification of further monogenic causes of nephrolithiasis are limited by the unavailability of large families To overcome these difficulties and facilitate the identification of genetic abnormalities causing idiopathic nephrolithiasis and nephrocalcinosis, we embarked on establishing mouse models for renal calcification by investigating the phenotypes of progeny of mice treated with the chemical mutagen N-ethyl-N-nitrosourea (ENU). We report an ENU-induced mouse model of renal calcification (RCALC), that is referred to as RCALC type 2 (RCALC2), and is associated with a Tyr265Stop (Y265X) nonsense mutation in the DNA polymerase subunit gamma-2, accessory protein (Polg2), which functions in the replication of mitochondrial DNA (mtDNA)encoded proteins
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