[show abstract][hide abstract] ABSTRACT: Mutations of PKD1 and PKD2 account for most cases of autosomal dominant polycystic kidney disease (ADPKD). Compared with PKD2, patients with PKD1 typically have more severe renal disease. Here, we report a follow-up study of a unique multigeneration family with bilineal ADPKD (NFL10) in which a PKD1 disease haplotype and a PKD2 (L736X) mutation co-segregated with 18 and 14 affected individuals, respectively. In our updated genotype-phenotype analysis of the family, we found that PKD1-affected individuals had uniformly mild renal disease similar to the PKD2-affected individuals. By sequencing all the exons and splice junctions of PKD1, we identified two missense mutations (Y528C and R1942H) from a PKD1-affected individual. Although both variants were predicted to be damaging to the mutant protein, only Y528C co-segregated with all of the PKD1-affected individuals in NFL10. Studies in MDCK cells stably expressing wild-type and mutant forms of PKD found that cell lines expressing the Y528C variant formed cysts in culture and displayed increased rates of growth and apoptosis. Thus, Y528C functions as a hypomorphic PKD1 allele. These findings have important implications for pathogenic mechanisms and molecular diagnostics of ADPKD.
Kidney International 02/2012; 81(4):412-7. · 7.92 Impact Factor
[show abstract][hide abstract] ABSTRACT: This article describes two sisters with type III Bartter syndrome (BS) due to a novel missense variant of the CLCNKB gene. The phenotypic expression of the disease was very different in these two siblings. In one sister, the disease followed a very severe course, especially in the neonatal period and as a toddler. Both the classic symptoms and the biochemical features of the syndrome were striking. In addition, she presented with sensorineural deafness, a complication yet unreported in this subtype of BS In contrast, the least affected sister was symptom free and the biochemical features of the disease although present remained discrete throughout the prolonged follow-up. It is suggested that such a difference in the phenotypic expression of the disease is possibly secondary to the modifier effect of a gene and/or results from environmental factor(s).
European Journal of Pediatrics 04/2011; 170(9):1209-11. · 1.91 Impact Factor
[show abstract][hide abstract] ABSTRACT: Mutations of PKD1 and PKD2 account for 85 and 15% of cases of autosomal dominant polycystic kidney disease (ADPKD), respectively. Clinically, PKD1 is more severe than PKD2, with a median age at ESRD of 53.4 versus 72.7 yr. In this study, we explored whether a family history of renal disease severity predicts the mutated gene in ADPKD. We examined the renal function (estimated GFR and age at ESRD) of 484 affected members from 90 families who had ADPKD and whose underlying genotype was known. We found that the presence of at least one affected family member who developed ESRD at age < or =55 was highly predictive of a PKD1 mutation (positive predictive value 100%; sensitivity 72%). In contrast, the presence of at least one affected family member who continued to have sufficient renal function or developed ESRD at age >70 was highly predictive of a PKD2 mutation (positive predictive value 100%; sensitivity 74%). These data suggest that close attention to the family history of renal disease severity in ADPKD may provide a simple means of predicting the mutated gene, which has prognostic implications.
Journal of the American Society of Nephrology 06/2009; 20(8):1833-8. · 8.99 Impact Factor
[show abstract][hide abstract] ABSTRACT: To elucidate the molecular pathways that modulate renal cyst growth in ADPKD, we performed global gene profiling on cysts of different size (<1 ml, n = 5; 10-20 ml, n = 5; >50 ml, n = 3) and minimally cystic tissue (MCT, n = 5) from five PKD1 human polycystic kidneys using Affymetrix HG-U133 Plus 2.0 arrays. We used gene set enrichment analysis to identify overrepresented signaling pathways and key transcription factors (TFs) between cysts and MCT. We found down-regulation of kidney epithelial restricted genes (e.g. nephron segment-specific markers and cilia-associated cystic genes such as HNF1B, PKHD1, IFT88 and CYS1) in the renal cysts. On the other hand, PKD1 cysts displayed a rich profile of gene sets associated with renal development, mitogen-mediated proliferation, cell cycle progression, epithelial-mesenchymal transition, hypoxia, aging and immune/inflammatory responses. Notably, our data suggest that up-regulation of Wnt/beta-catenin, pleiotropic growth factor/receptor tyrosine kinase (e.g. IGF/IGF1R, FGF/FGFR, EGF/EGFR, VEGF/VEGFR), G-protein-coupled receptor (e.g. PTGER2) signaling was associated with renal cystic growth. By integrating these pathways with a number of dysregulated networks of TFs (e.g. SRF, MYC, E2F1, CREB1, LEF1, TCF7, HNF1B/ HNF1A and HNF4A), our data suggest that epithelial dedifferentiation accompanied by aberrant activation and cross-talk of specific signaling pathways may be required for PKD1 cyst growth and disease progression. Pharmacological modulation of some of these signaling pathways may provide a potential therapeutic strategy for ADPKD.
Human Molecular Genetics 04/2009; 18(13):2328-43. · 7.69 Impact Factor
[show abstract][hide abstract] ABSTRACT: Mutation-based molecular diagnostics of autosomal dominant polycystic kidney disease (ADPKD) is complicated by locus and allelic heterogeneity, large multi-exon gene structure and duplication in PKD1, and a high level of unclassified variants. Comprehensive screening of PKD1 and PKD2 by two recent studies have shown that atypical splice mutations account for 3.5% to 5% of ADPKD. We evaluated the role of bioinformatic prediction of atypical splice mutations and determined the pathogenicity of an atypical PKD2 splice variant from a multiplex ADPKD (TOR101) family.
Using PubMed, we identified 17 atypical PKD1 and PKD2 splice mutations. We found that bioinformatics analysis was often useful for evaluating the pathogenicity of these mutations, although RT-PCR is needed to provide the definitive proof.
Sequencing of both PKD1 and PKD2 in an affected subject of TOR101 failed to identify a definite mutation, but revealed several UCVs, including an atypical PKD2 splice variant. Linkage analysis with microsatellite markers indicated that TOR101 was PKD2-linked and IVS8 + 5G-->A was shown to cosegregate only with affected subjects. RT-PCR of leukocyte mRNA from an affected subject using primers from exons 7 and 9 revealed six splice variants that resulted from activation of different combinations of donor and acceptor cryptic splice sites, all terminating with premature stop codons.
The data provide strong evidence that IVS8 + 5G-->A is a pathogenic mutation for PKD2. This case highlights the importance of functional analysis of UCVs.
Clinical Journal of the American Society of Nephrology 01/2009; 4(2):442-9. · 5.07 Impact Factor
[show abstract][hide abstract] ABSTRACT: Familial clustering and genome-wide linkage scans strongly support a genetic susceptibility to familial IgA nephropathy (IgAN), but genetic factors that predispose to sporadic IgAN are unknown. A high-throughput single nucleotide polymorphism (SNP) association study was conducted using a customized Illumina BeadChip in 732 white patients with biopsy-proven IgAN and 503 control subjects from Canada, France, and Finland. Approximately 93% of 1536 SNPs on the array were tag SNPs from Phase I+II of the HapMap with a minor allele frequency > or =5%, designed to capture the common variants of genes within the critical interval of IGAN1 on chromosome 6q22 and 69 biologic candidate genes for IgAN. SNPs of suggestive or significant association were identified by using logistic regression to adjust for age, gender, study site, and population stratification. Despite using a dense marker set that covered an average interval of 6.5 kb between SNPs, there was no strong and consistent association signal within the IGAN1 critical interval. Among the biologic candidate genes examined, two significant association signals were found at IL5RA and TNFRSF6B, the latter being particularly interesting because this gene encodes a decoy receptor for a TNF family ligand that causes IgAN in mice when overexpressed. Pending replication, these data suggest that variants of IL5RA and TNFRSF6B may predispose to sporadic IgAN.
Journal of the American Society of Nephrology 06/2008; 19(5):1025-33. · 8.99 Impact Factor
[show abstract][hide abstract] ABSTRACT: Gene-based mutation screening is now available and has the potential to provide diagnostic confirmation or exclusion of autosomal dominant polycystic kidney disease. This study illustrates its utility and limitations in the clinical setting.
Using a molecular diagnostic service, genomic DNA of one affected individual from each study family was screened for pathologic PKD1 and PKD2 mutations. Bidirectional sequencing was performed to identify sequence variants in all exons and splice junctions of both genes and to confirm the specific mutations in other family members. In two multiplex families, microsatellite markers were genotyped at both PDK1 and PKD2 loci, and pair-wise and multipoint linkage analysis was performed.
Three of five probands studied were referred for assessment of renal cystic disease without a family history of autosomal dominant polycystic kidney disease, and two others were younger at-risk members of families with autosomal dominant polycystic kidney disease being evaluated as living-related kidney donors. Gene-based mutation screening identified pathogenic mutations that provided confirmation or exclusion of disease in three probands, but in the other two, only unclassified variants were identified. In one proband in which mutation screening was indeterminate, DNA linkage studies provided strong evidence for disease exclusion.
Gene-based mutation screening or DNA linkage analysis should be considered in individuals in whom the diagnosis of autosomal dominant polycystic kidney disease is uncertain because of a lack of family history or equivocal imaging results and in younger at-risk individuals who are being evaluated as living-related kidney donors.
Clinical Journal of the American Society of Nephrology 02/2008; 3(1):146-52. · 5.07 Impact Factor
[show abstract][hide abstract] ABSTRACT: Recessive NPHS2 (podocin) mutations account for up to approximately 30% of steroid-resistant idiopathic FSGS in children and are associated with a reduced risk for disease recurrence after renal transplantation. R229Q, a missense variant that is present in 3.6% of the white population, has been implicated as a common disease-causing mutation. Given these clinical implications, we examined the role of NPHS2 mutations in a cohort of patients with adult-onset FSGS. We used denaturing HPLC to screen for heterozygous and homozygous gene variants in PCR-amplified DNA fragments that contained all exons and splice junctions of NPHS2. Bidirectional sequencing was performed to define all of the gene variants detected. With the use of the denaturing HPLC in a single-blind pilot study, 40 of 43 known NPHS2 mutations were detected from 22 pediatric patients with FSGS to establish a test sensitivity of 93%. This screen then was applied to 87 adult patients with idiopathic FSGS (15 steroid-sensitive, 63 steroid-resistant, and nine familial cases). In this latter cohort, compound heterozygous mutations were detected only in one patient with steroid-sensitive FSGS (R229Q and Q285fsX302) and no homozygous mutations. Overall, R229Q accounted for eight (80%) of ten of the putative mutant alleles that were detected in the study cohort. Contrary to the pediatric experience, recessive NPHS2 mutations are rare in this study population, suggesting that the pathogenesis of FSGS in adults may differ from that in children. These data do not support R229Q as a disease-causing mutation for steroid-resistant FSGS.
Clinical Journal of the American Society of Nephrology 02/2007; 2(1):31-7. · 5.07 Impact Factor
[show abstract][hide abstract] ABSTRACT: The hypoparathyroidism, deafness, and renal dysplasia (HDR) syndrome is an autosomal dominant disorder caused by mutations of a member of the GATA-binding family of transcription factors, GATA3. This dual zinc finger transcription factor binds DNA with its C-terminal zinc finger (ZnF2) and stabilizes this binding with its N-terminal zinc finger (ZnF1). ZnF1 also interacts with other zinc finger proteins, notably Friend of GATA (FOG). The HDR syndrome has been described in patients with mutations affecting both ZnF1 and ZnF2 domains; the former result in inefficient interaction with FOG, and the latter result in disruption of DNA binding. We report a patient with renal failure, hypoparathyroidism, and bilateral hearing loss. Assessment of family members indicated that the disease arose as a de novo mutation in her mother. Analysis of GATA3 in the family revealed a heterozygous missense mutation resulting in a nonconservative change of a single amino acid (R276P) in the ZnF1 domain. Functional analysis using dissociation electrophoretic mobility shift and yeast two-hybrid assays showed reduced binding affinity to the GATA motifs but normal interaction with FOG in vitro. These results are consistent with the predicted functions of human GATA3-ZnF1 from three-dimensional molecular modeling and with HDR being a result of GATA3 haploinsufficiency.
[show abstract][hide abstract] ABSTRACT: Significant intrafamilial phenotypic variability is well documented in autosomal dominant polycystic kidney disease (ADPKD) and suggests a modifier effect. In this study, variance components analysis was performed to estimate the contribution of genetic factors for within-family renal disease variability in 406 patients from 66 type 1 ADPKD families. Overall, 39% of the study patients had ESRD at their last follow-up, and their renal survival did not differ by gender (P = 0.35, log-rank test). Because their frequency plot of creatinine clearance (Ccr) assumed a bimodal distribution with a marked kurtosis that was not improved by transformations, the study cohort was decomposed into two separate groups (non-ESRD [n = 247] and ESRD [n = 159]) in which the Ccr plots were normally distributed. The heritability (h(2)) of Ccr and age at ESRD (age(ESRD)) and the genetic correlations between these measures and their covariates were estimated. In patients without ESRD, a significant heritability was found for Ccr (h(2) = 0.42; P = 0.0015) after adjusting for age (P = 0.0001), systolic BP (P = 0.0006), and treatment with angiotensin-converting enzyme inhibitor/angiotensin receptor blocker (P = 0.00001). Birth year, gender, BMI, diastolic and mean BP, and pack-years of cigarette smoking did not significantly influence the heritability of this trait. In patients with ESRD, age(ESRD) provides a better measure than Ccr, which was very narrowly distributed. A significant heritability was found for age(ESRD) (h(2) = 0.78; P = 0.00009) in these latter patients. None of the above covariates influenced the heritability of this trait. It is concluded that a significant modifier gene effect influences the progression of renal disease in type 1 ADPKD.
Journal of the American Society of Nephrology 04/2005; 16(3):755-62. · 8.99 Impact Factor
[show abstract][hide abstract] ABSTRACT: Autosomal dominant polycystic kidney disease (ADPKD) is a common Mendelian disorder that affects approximately 1 in 1000 live births. Mutations of two genes, PKD1 and PKD2, account for the disease in approximately 80 to 85% and 10 to 15% of the cases, respectively. Significant interfamilial and intrafamilial renal disease variability in ADPKD has been well documented. Locus heterogeneity is a major determinant for interfamilial disease variability (i.e., patients from PKD1-linked families have a significantly earlier onset of ESRD compared with patients from PKD2-linked families). More recently, two studies have suggested that allelic heterogeneity might influence renal disease severity. The current study examined the genotype-renal function correlation in 461 affected individuals from 71 ADPKD families with known PKD2 mutations. Fifty different mutations were identified in these families, spanning between exon 1 and 14 of PKD2. Most (94%) of these mutations were predicted to be inactivating. The renal outcomes of these patients, including the age of onset of end-stage renal disease (ESRD) and chronic renal failure (CRF; defined as creatinine clearance < or = 50 ml/min, calculated using the Cockroft and Gault formula), were analyzed. Of all the affected individuals clinically assessed, 117 (25.4%) had ESRD, 47 (10.2%) died without ESRD, 65 (14.0%) had CRF, and 232 (50.3%) had neither CRF nor ESRD at the last follow-up. Female patients, compared with male patients, had a later mean age of onset of ESRD (76.0 [95% CI, 73.8 to 78.1] versus 68.1 [95% CI, 66.0 to 70.2] yr) and CRF (72.5 [95% CI, 70.1 to 74.9] versus 63.7 [95% CI, 61.4 to 66.0] yr). Linear regression and renal survival analyses revealed that the location of PKD2 mutations did not influence the age of onset of ESRD. However, patients with splice site mutations appeared to have milder renal disease compared with patients with other mutation types (P < 0.04 by log rank test; adjusted for the gender effect). Considerable renal disease variability was also found among affected individuals with the same PKD2 mutations. This variability can confound the determination of allelic effects and supports the notion that additional genetic and/or environmental factors may modulate the renal disease severity in ADPKD.
Journal of the American Society of Nephrology 05/2003; 14(5):1164-74. · 8.99 Impact Factor
[show abstract][hide abstract] ABSTRACT: In searching for a putative third gene for autosomal dominant polycystic kidney disease (ADPKD), we studied the genetic inheritance of a large family (NFL10) previously excluded from linkage to both the PKD1 locus and the PKD2 locus. We screened 48 members of the NFL10 pedigree, by ultrasonography, and genotyped them, with informative markers, at both the PKD1 locus and the PKD2 locus. Twenty-eight of 48 individuals assessed were affected with ADPKD. Inspection of the haplotypes of these individuals suggested the possibility of bilineal disease from independently segregating PKD1 and PKD2 mutations. Using single-stranded conformational analysis, we screened for and found a PKD2 mutation (i.e., 2152delA; L736X) in 12 affected pedigree members. Additionally, when the disease status of these individuals was coded as "unknown" in linkage analysis, we also found, with markers at the PKD1 locus, significant LOD scores (i.e., >3.0). These findings strongly support the presence of a PKD1 mutation in 15 other affected pedigree members, who lack the PKD2 mutation. Two additional affected individuals had trans-heterozygous mutations involving both genes, and they had renal disease that was more severe than that in affected individuals who had either mutation alone. This is the first documentation of bilineal disease in ADPKD. In humans, trans-heterozygous mutations involving both PKD1 and PKD2 are not necessarily embryonically lethal. However, the disease associated with the presence of both mutations appears to be more severe than the disease associated with either mutation alone. The presence of bilineal disease as a confounder needs to be considered seriously in the search for the elusive PKD3 locus.
The American Journal of Human Genetics 02/2001; 68(2):355-63. · 11.20 Impact Factor
[show abstract][hide abstract] ABSTRACT: Autosomal dominant polycystic kidney disease (ADPKD) is caused by mutations in PKD1 and PKD2. The products of these genes associate to form heteromeric complexes. Several models have been proposed to explain the mechanism of cyst formation. Here we find somatic mutations of PKD2 in 71% of ADPKD2 cysts analysed. Clonal somatic mutations of PKD1 were identified in a subset of cysts that lacked PKD2 mutations.
[show abstract][hide abstract] ABSTRACT: An intriguing feature of autosomal dominant polycystic kidney disease (ADPKD) is the focal and sporadic formation of renal and extrarenal cysts. Recent documentation of somatic PKD1 mutations in cystic epithelia of patients with germ-line PKD1 mutations suggests a "two-hit" model for cystogenesis in type 1 ADPKD. This study tests whether the same mechanism for cystogenesis might also occur in type 2 ADPKD. Genomic DNA was obtained from 54 kidney and liver cysts from three patients with known germ-line PKD2 mutations, using procedures that minimize contamination of cells from noncystic tissue. Using intragenic and microsatellite markers, these cyst samples were screened for loss of heterozygosity. The same samples were also screened for somatic mutations in five of the 15 exons in PKD2 by single-stranded conformational polymorphism analysis. Loss of heterozygosity was found in five cysts, and unique intragenic mutations were found in seven other cysts. In 11 of these 12 cysts, it was also determined that the somatic mutation occurred nonrandomly in the copy of PKD2 inherited from the unaffected parent. These findings support the "two-hit" model as a unified mechanism for cystogenesis in ADPKD. In this model, the requirement of a somatic mutation as the rate-limiting step for individual cyst formation has potential therapeutic implications.
Journal of the American Society of Nephrology 08/1999; 10(7):1524-9. · 8.99 Impact Factor
[show abstract][hide abstract] ABSTRACT: Autosomal dominant polycystic kidney disease (ADPKD) is a common Mendelian disorder that affects approximately 1 in 1000 live births. Linkage studies have shown that the majority (approximately 85%) of cases are due to mutations in PKD1 on chromosome 16p, while mutations in PKD2 on chromosome 4q account for most of the remaining cases. Locus heterogeneity in ADPKD is known to contribute to differences in disease severity, with PKD1-linked families having earlier onset of end-stage renal disease (ESRD) than PKD2-linked families (mean age at ESRD: 56 versus 70, respectively). In this study, 11 Canadian families with ADPKD were screened for PKD2 mutations. In four families, linkage to PKD2 was previously documented. In the remaining seven smaller families, one or more affected members had late-onset ESRD at age 70 or older. Using single-stranded conformational polymorphism analysis, one affected member from each family was screened for mutations in all 15 exons of PKD2, which were PCR-amplified from genomic templates. A spectrum of mutations was found in approximately 73% (8 of 11) of the families screened, with no difference in the detection rate between the PKD2-linked families and the families with late-onset ESRD. In three unrelated families, insertion or deletion of an adenosine in a polyadenosine tract (i.e., (A)8 at nt 2152-2159) was found on exon 11, suggesting that this mononucleotide repeat tract is prone to mutations from "slipped strand mispairing." All mutations, scattered between exons 1 and 11, are predicted to result in a truncated polycystin 2 that lacks both the calcium-binding EF-hand domain and the two cytoplasmic domains required for the interaction of polycystin 2 with polycystin 1 and with itself. Furthermore, no correlation was found between the location of the mutations in the PKD2 coding sequence and disease severity. Thus, these findings are consistent with other recently published reports and suggest that most PKD2 mutations are inactivating.
Journal of the American Society of Nephrology 11/1998; 9(10):1853-60. · 8.99 Impact Factor