Linkage confirms canine pkd1 orthologue as a candidate for bull terrier polycystic kidney disease

Bull terrier polycystic kidney disease (BTPKD) is a Mendelian disorder with many features reminiscent of human autosomal dominant polycystic disease, the latter disease being due to mutations at PKD1 and PKD2 loci. We investigated the role of the canine pkd1 orthologue in BTPKD via linkage analysis of a large kindred in which the disorder is segregating. Twelve microsatellite markers around the canine pkd1 locus (CFA6) were amplified from the genomic DNA of 20 affected and 16 unaffected bull terriers. An additional 28 affected dogs were genotyped at five key microsatellites. A highly significant multi-point LOD score that peaked over the canine pkd1 locus was observed (LOD = 6.59, best two-point LOD score LOD = 6.02), implicating this as the BTPKD locus.     

Molecular and cellular pathophysiology of autosomal recessive polycystic kidney disease (ARPKD)

Autosomal recessive polycystic kidney disease (ARPKD) belongs to a group of congenital hepatorenal fibrocystic syndromes characterized by dual renal and hepatic involvement of variable severity. Despite the wide clinical spectrum of ARPKD (MIM 263200), genetic linkage studies indicate that mutations at a single locus, PKHD1 (polycystic kidney and hepatic disease 1), located on human chromosome region 6p21.1–p12, are responsible for all phenotypes of ARPKD. Identification of cystic disease genes and their encoded proteins has provided investigators with critical tools to begin to unravel the molecular and cellular mechanisms of PKD. PKD cystic epithelia share common phenotypic abnormalities despite the different genetic mutations that underlie the disease. Recent studies have shown that many cyst-causing proteins are expressed in multimeric complexes at distinct subcellular locations within epithelia. This co-expression of cystoproteins suggests that cyst formation, regardless of the underlying disease gene, results from perturbations in convergent and/or integrated signal transduction pathways. To date, no specific therapies are in clinical use for ameliorating cyst growth in ARPKD. However, studies noted in this review suggest that therapeutic targeting of the cAMP and epidermal growth factor receptor (EGFR)-axis abnormalities in cystic epithelia may translate into effective therapies for ARPKD and, by analogy, autosomal dominant polycystic kidney disease (ADPKD). A particularly promising approach appears to be the targeting of downstream intermediates of both the cAMP and EGFR axis. This review focuses on ARPKD and presents a concise summary of the current understanding of the molecular genetics and cellular pathophysiology of this disease. It also highlights phenotypic and mechanistic similarities between ARPKD and ADPKD.The authors are supported by the National Institutes of Health (grant no. 1-P50-DK57306), the PKD Foundation (grant no. 76a2r), and the Children’s Research Institute, Children’s Hospital of Wisconsin.     

Cilia and polycystic kidney disease,kith and kin

In the past decade, cilia have been found to play important roles in renal cystogenesis. Many genes, such as PKD1 and PKD2 which, when mutated, cause autosomal dominant polycystic kidney disease (ADPKD), have been found to localize to primary cilia. The cilium functions as a sensor to transmit extracellular signals into the cell. Abnormal cilia structure and function are associated with the development of polyscystic kidney disease (PKD). Cilia assembly includes centriole migration to the apical surface of the cell, ciliary vesicle docking and fusion with the cell membrane at the intended site of cilium outgrowth, and microtubule growth from the basal body. This review summarizes the most recent advances in cilia and PKD research, with special emphasis on the mechanisms of cytoplasmic and intraciliary protein transport during ciliogenesis. Birth Defects Research (Part C) 102:174–185, 2014 . © 2014 Wiley Periodicals, Inc .     

A non-synonymous mutation in the canine Pkd1 gene is associated with autosomal dominant polycystic kidney disease in Bull Terriers

Polycystic Kidney Disease is an autosomal dominant disease common in some lines of Bull Terriers (BTPKD). The disease is linked to the canine orthologue of human PKD1 gene, Pkd1, located on CFA06, but no disease-associated mutation has been reported. This study sequenced genomic DNA from two Bull Terriers with BTPKD and two without the disease. A non-synonymous G>A transition mutation in exon 29 of Pkd1 was identified. A TaqMan® SNP Genotyping Assay was designed and demonstrated the heterozygous detection of the mutation in 47 Bull Terriers with BTPKD, but not in 102 Bull Terriers over one year of age and without BTPKD. This missense mutation replaces a glutamic acid residue with a lysine residue in the predicted protein, Polycystin 1. This region of Polycystin 1 is highly conserved between species, and is located in the first cytoplasmic loop of the predicted protein structure, close to the PLAT domain and the second transmembrane region. Thus, this change could alter Polycystin 1 binding or localization. Analytic programs PolyPhen 2, Align GVGD and SIFT predict this mutation to be pathogenic. Thus, BTPKD is associated with a missense mutation in Pkd1, and the application of this mutation specific assay could reduce disease transmission by allowing diagnosis of disease in young animals prior to breeding.     

Role of the N-terminal polycystic kidney disease domain in chitin degradation by chitinase A from a marine bacterium, Alteromonas sp. strain O-7

AIMS: The aim of study was to clarify whether the polycystic kidney disease (PKD) domain of chitinase A (ChiA) participates in the hydrolysis of powdered chitin. METHODS AND RESULTS: Site-directed mutagenesis of the conserved aromatic residues of PKD domain was performed by PCR. The aromatic residues, W30, Y48, W64 and W67, were replaced by alanine, and single- and double-mutant chitinases were produced in Escherichia coli XL10 and purified with HisTrap column. Single mutations were not quite effective on the hydrolysing activities against chitinous substrates when compared with wild-type ChiA. However, mutations of W30 and W67 decreased the activities against powdered chitin by 87.6%. Wild-type and mutant PKD domains were produced in E. coli TOP10 and purified with glutathione-Sepharose 4B column. Wild-type PKD domain showed significant binding activity to powdered chitin, whereas mutations of W30 and W67 reduced the binding activity to powdered chitin drastically. These results suggest that PKD domain of ChiA is essential for effective hydrolysis of powdered chitin through the interaction between two aromatic residues and chitin molecule. CONCLUSIONS: PKD domain of ChiA participates in the effective hydrolysis of powdered chitin through the interaction between two aromatic residues (W30 and W67) and chitin molecule. SIGNIFICANCE AND IMPACT OF THE STUDY: The findings of this study provide important information on chitin degradation by microbial chitinases.     

Regional localization of the autosomal dominant polycystic kidney disease locus

The localization of the autosomal dominant polycystic kidney disease locus (PKD1) within an array of anonymous polymorphic DNA sequences on chromosome 16 band p13 was determined by multipoint mapping. Nine polymorphic DNA markers, including two hypervariable sequences, were used to study 19 PKD1 and 21 reference families. PKD1 was found to lie proximal to the 3' and 5' hypervariable regions of alpha-globin and distal to the anonymous sequence CRI-0327. Somatic cell hybrid mapping places PKD1 within the region 16p13.11-16pter. The availability of an array of linked markers which bracket the PKD1 locus provides a framework for further attempts to identify the PKD1 gene and offers an improved method of presymptomatic diagnosis of the disease.     

Autosomal dominant polycystic kidney disease: Disrupted pathways and potential therapeutic interventions

Autosomal dominant polycystic kidney disease (ADPKD) is a monogenic inherited renal cystic disease that occurs in different races worldwide. It is characterized by the development of a multitude of renal cysts, which leads to massive enlargement of the kidney and often to renal failure in adulthood. ADPKD is caused by a mutation in PKD1 or PKD2 genes encoding the proteins polycystin-1 and polycystin-2, respectively. Recent studies showed that cyst formation and growth result from deregulation of multiple cellular pathways like proliferation, apoptosis, metabolic processes, cell polarity, and immune defense. In ADPKD, intracellular cyclic adenosine monophosphate (cAMP) promotes cyst enlargement by stimulating cell proliferation and transepithelial fluid secretion. Several interventions affecting many of these defective signaling pathways have been effective in animal models and some are currently being tested in clinical trials. Moreover, the stem cell therapy can improve nephropathies and according to studies were done in this field, can be considered as a hopeful therapeutic approach in future for PKD. This study provides an in-depth review of the relevant molecular pathways associated with the pathogenesis of ADPKD and their implications in development of potential therapeutic strategies.     

Comprehensive strategy improves the genetic diagnosis of different polycystic kidney diseases

Polycystic kidney disease (PKD) is known to occur in three main forms, namely autosomal dominant PKD (ADPKD), autosomal recessive PKD (ARPKD) and syndromic PKD (SPKD), based on the clinical manifestations and genetic causes, which are diagnosable from the embryo stage to the later stages of life. Selection of the genetic test for the individuals with diagnostic imaging reports of cystic kidneys without a family history of the disease continues to be a challenge in clinical practice. With the objective of maintaining a limit on the time and medical cost of the procedure, a practical strategy for genotyping and targeted validation to resolve cystogene variations was developed in our clinical laboratory, which combined the techniques of whole-exome sequencing (WES), Long-range PCR (LR-PCR), Sanger sequencing and multiplex ligation–dependent probe amplification (MLPA) to work in a stepwise approach. In this context, twenty-six families with renal polycystic disorders were enrolled in the present study. Thirty-two variants involving four ciliary genes (PKD1, PKHD1, TMEM67 and TMEM107) were identified and verified in 23 families (88.5%, 23/26), which expanded the variant spectrum by 16 novel variants. Pathogenic variations in five foetuses of six families diagnosed with PKD were identified using prenatal ultrasound imaging. Constitutional biallelic and digenic variations constituted the pathogenic patterns in these foetuses. The preliminary clinical data highlighted that the WES + LR PCR-based workflow followed in the present study is efficient in detecting divergent variations in PKD. The biallelic and digenic mutations were revealed as the main pathogenic patterns in the foetuses with PKD.     

Novel mutations of the PKD1 gene in Korean patients with autosomal dominant polycystic kidney disease

The gene for the most common form of autosomal dominant polycystic kidney disease (ADPKD), PKD1, has recently been characterized and shown to encode an integral membrane protein, polycystin-1, which is involved in cell-cell and cell-matrix interactions. Until now, approximately 30 mutations of the 3' single copy region of the PKD1 gene have been reported in European and American populations. However, there is no report of mutations in Asian populations. Using the polymerase chain reaction and single-strand conformation polymorphism (SSCP) analysis, 91 Korean patients with ADPKD were screened for mutation in the 3' single copy region of the PKD1 gene. As a result, we have identified and characterized six mutations: three frameshift mutations (11548del8bp, 11674insG and 12722delT), a nonsense mutation (Q4010X), and two missense mutations (R3752W and D3814N). Five mutations except for Q4010X are reported here for the first time. Our findings also indicate that many different mutations are likely to be responsible for ADPKD in the Korean population. The detection of additional disease-causing PKD1 mutations will help in identifying the location of the important functional regions of polycystin-1 and help us to better understand the pathophysiology of ADPKD.     

Renal cystic disease in tuberous sclerosis: role of the polycystic kidney disease 1 gene.

Tuberous sclerosis is an autosomal dominant trait characterized by the development of hamartomatous growths in many organs. Renal cysts are also a frequent manifestation. Major genes for tuberous sclerosis and autosomal dominant polycystic kidney disease, TSC2 and PKD1, respectively, lie adjacent to each other at chromosome 16p13.3, suggesting a role for PKD1 in the etiology of renal cystic disease in tuberous sclerosis. We studied 27 unrelated patients with tuberous sclerosis and renal cystic disease. Clinical histories and radiographic features were reviewed, and renal function was assessed. We sought mutations at the TSC2 and PKD1 loci, using pulsed field- and conventional-gel electrophoresis and FISH. Twenty-two patients had contiguous deletions of TSC2 and PKD1. In 17 patients with constitutional deletions, cystic disease was severe, with early renal insufficiency. One patient with deletion of TSC2 and of only the 3' UTR of PKD1 had few cysts. Four patients were somatic mosaics; the severity of their cystic disease varied considerably. Mosaicism and mild cystic disease also were demonstrated in parents of 3 of the constitutionally deleted patients. Five patients without contiguous deletions had relatively mild cystic disease, 3 of whom had gross rearrangements of TSC2 and 2 in whom no mutation was identified. Significant renal cystic disease in tuberous sclerosis usually reflects mutational involvement of the PKD1 gene, and mosaicism for large deletions of TSC2 and PKD1 is a frequent phenomenon.     

Bilineal disease and trans-heterozygotes in autosomal dominant polycystic kidney disease

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 cell biology of polycystic kidney disease

Polycystic kidney disease is a common genetic disorder in which fluid-filled cysts displace normal renal tubules. Here we focus on autosomal dominant polycystic kidney disease, which is attributable to mutations in the PKD1 and PKD2 genes and which is characterized by perturbations of renal epithelial cell growth control, fluid transport, and morphogenesis. The mechanisms that connect the underlying genetic defects to disease pathogenesis are poorly understood, but their exploration is shedding new light on interesting cell biological processes and suggesting novel therapeutic targets.     

Molecular pathogenesis of autosomal dominant polycystic kidney disease

Autosomal dominant polycystic kidney disease (ADPKD) is one of the commonest inherited human disorders yet remains relatively unknown to the wider medical, scientific and public audience. ADPKD is characterised by the development of bilateral enlarged kidneys containing multiple fluid-filled cysts and is a leading cause of end-stage renal failure (ESRF). ADPKD is caused by mutations in two genes: PKD1 and PKD2. The protein products of the PKD genes, polycystin-1 and polycystin-2, form a calcium-regulated, calcium-permeable ion channel. The polycystin complex is implicated in regulation of the cell cycle via multiple signal transduction pathways as well as the mechanosensory function of the renal primary cilium, an enigmatic cellular organelle whose role in normal physiology is still poorly understood. Defects in cilial function are now documented in several other human diseases including autosomal recessive polycystic kidney disease, nephronophthisis, Bardet-Biedl syndrome and many animal models of polycystic kidney disease. Therapeutic trials in these animal models of polycystic kidney disease have identified several promising drugs that ameliorate disease severity. However, elucidation of the function of the polycystins and the primary cilium will have a major impact on our understanding of renal cystic diseases and will create exciting new opportunities for the design of disease-specific therapies.     

Evidence of linkage disequilibrium in the Spanish polycystic kidney disease I population.

Forty-one Spanish families with polycystic kidney disease 1 (PKD1) were studied for evidence of linkage disequilibrium between the disease locus and six closely linked markers. Four of these loci--three highly polymorphic microsatellites (SM6, CW3, and CW2) and an RFLP marker (BLu24)--are described for the first time in this report. Overall the results reveal many different haplotypes on the disease-carrying chromosome, suggesting a variety of independent PKD1 mutations. However, linkage disequilibrium was found between BLu24 and PKD1, and this was corroborated by haplotype analysis including the microsatellite polymorphisms. From this analysis a group of closely related haplotypes, consisting of four markers, was found on 40% of PKD1 chromosomes, although markers flanking this homogeneous region showed greater variability. This study has highlighted an interesting subpopulation of Spanish PKD1 chromosomes, many of which have a common origin, that may be useful for localizing the PKD1 locus more precisely.     

Mouse models of polycystic kidney disease induced by defects of ciliary proteins

Polycystic kidney disease (PKD) is a common hereditary disorder which is characterized by fluid-filled cysts in the kidney. Mutation in either PKD1, encoding polycystin-1 (PC1), or PKD2, encoding polycystin-2 (PC2), are causative genes of PKD. Recent studies indicate that renal cilia, known as mechanosensors, detecting flow stimulation through renal tubules, have a critical function in maintaining homeostasis of renal epithelial cells. Because most proteins related to PKD are localized to renal cilia or have a function in ciliogenesis. PC1/PC2 heterodimer is localized to the cilia, playing a role in calcium channels. Also, disruptions of ciliary proteins, except for PC1 and PC2, could be involved in the induction of polycystic kidney disease. Based on these findings, various PKD mice models were produced to understand the roles of primary cilia defects in renal cyst formation. In this review, we will describe the general role of cilia in renal epithelial cells, and the relationship between ciliary defects and PKD. We also discuss mouse models of PKD related to ciliary defects based on recent studies. [BMB Reports 2013; 46(2): 73-79]     

Apoptosis in polycystic kidney disease

Apoptosis is the process of programmed cell death. It is a ubiquitous, controlled process consuming cellular energy and designed to avoid cytokine release despite activation of local immune cells, which clear the cell fragments. The process occurs during organ development and in maintenance of homeostasis. Abnormalities in any step of the apoptotic process are associated with autoimmune diseases and malignancies. Polycystic kidney disease (PKD) is the most common inherited kidney disease leading to end-stage renal disease (ESRD). Cyst formation requires multiple mechanisms and apoptosis is considered one of them. Abnormalities in apoptotic processes have been described in various murine and rodent models of PKD as well as in human PKD kidneys. The purpose of this review is to outline the role of apoptosis in progression of PKD as well as to describe the mechanisms involved. This article is part of a Special Issue entitled: Polycystic Kidney Disease.     

A loss-of-function model for cystogenesis in human autosomal dominant polycystic kidney disease type 2.

Autosomal dominant polycystic kidney disease (ADPKD) is genetically heterogeneous, with at least three chromosomal loci (PKD1, PKD2, and PKD3) that account for the disease. Mutations in the PKD2 gene, on the long arm of chromosome 4, are expected to be responsible for approximately 15% of cases of ADPKD. Although ADPKD is a systemic disease, it shows a focal expression, because <1% of nephrons become cystic. A feasible explanation for the focal nature of events in PKD1, proposed on the basis of the two-hit theory, suggests that cystogenesis results from the inactivation of the normal copy of the PKD1 gene by a second somatic mutation. The aim of this study is to demonstrate that somatic mutations are present in renal cysts from a PKD2 kidney. We have studied 30 renal cysts from a patient with PKD2 in which the germline mutation was shown to be a deletion that encompassed most of the disease gene. Loss-of-heterozygosity (LOH) studies showed loss of the wild-type allele in 10% of cysts. Screening of six exons of the gene by SSCP detected eight different somatic mutations, all of them expected to produce truncated proteins. Overall, >/=37% of the cysts studied presented somatic mutations. No LOH for the PKD1 gene or locus D3S1478 were observed in those cysts, which demonstrates that somatic alterations are specific. We have identified second-hit mutations in human PKD2 cysts, which suggests that this mechanism could be a crucial event in the development of cystogenesis in human ADPKD-type 2.     

Autosomal dominant polycystic kidney disease: Genetics, mutations and microRNAs

Autosomal dominant polycystic kidney disease (ADPKD) is a common, monogenic multi-systemic disorder characterized by the development of renal cysts and various extrarenal manifestations. Worldwide, it is a common cause of end-stage renal disease. ADPKD is caused by mutation in either one of two principal genes, PKD1 and PKD2, but has large phenotypic variability among affected individuals, attributable to PKD genic and allelic variability and, possibly, modifier gene effects. Recent studies have generated considerable information regarding the genetic basis and molecular diagnosis of this disease, its pathogenesis, and potential strategies for targeted treatment. The purpose of this article is to provide a comprehensive review of the genetics of ADPKD, including mechanisms responsible for disease development, the role of gene variations and mutations in disease presentation, and the putative role of microRNAs in ADPKD etiology. The emerging and important role of genetic testing and the advent of novel molecular diagnostic applications also are reviewed. This article is part of a Special Issue entitled: Polycystic Kidney Disease.     

Animal models of polycystic kidney disease

Polycystic kidney disease (PKD) is one of the most prevalent causes of heritable renal failure. The disease is characterized by the occurrence of numerous fluid-filled cysts within the parenchyma of kidney. The cysts are epithelial in origin and expand in size, leading to crowding of normal kidney tissue. Ultimately, there is gross enlargement of the kidneys with loss of normal functions, and death usually occurs because of complications related to renal failure. Animal models of polycystic kidney disease are proving to be extremely useful for studying the molecular basis of renal cyst formation and for the isolation of genes carrying the mutations. This article describes the various animal models of polycystic kidney disease, spontaneously and experimentally derived, that have recently been identified.