Search

Digenic inheritance (DI) concerns pathologies with the simplest form of multigenic etiology, implicating more than 1 gene (and perhaps the environment). True DI is when biallelic or even triallelic mutations in 2 distinct genes, in cis or in trans, are necessary and sufficient to cause pathology with a defined diagnosis. In true DI, a heterozygous mutation in each of 2 genes alone is not associated with a recognizable phenotype. Well‐documented diseases with true DI are so far rare and follow non‐Mendelian inheritance. DI is also encountered when by serendipity, pathogenic mutations responsible for 2 distinct disease entities are co‐inherited, leading to a mixed phenotype. Also, we can consider many true monogenic Mendelian conditions, which show impressively broad spectrum of phenotypes due to pseudo‐DI, as a result of co‐inheriting genetic modifiers (GMs). I am herewith reviewing examples of GM and embark on presenting some recent notable examples of true DI, with wider discussion of the literature. Undeniably, the advent of high throughput sequencing is bound to unravel more patients suffering with true DI conditions and elucidate many important GM, thus impacting precision medicine. [ABSTRACT FROM AUTHOR]

Demosthenous P, Voskarides K, Stylianou K, Hadjigavriel M, Arsali M, Patsias C, Georgaki E, Zirogiannis P, Stavrou C, Daphnis E, Pierides A, Deltas C, Hellenic Nephrogenetics Research Consortium. X-linked Alport syndrome in Hellenic families: Phenotypic heterogeneity and mutations near interruptions of the collagen domain in COL4A5. The X-linked Alport syndrome (ATS) is caused by mutations in COL4A5 and exhibits a widely variable expression. Usually ATS is heralded with continuous microhematuria which rapidly progresses to proteinuria, hypertension and chronic or end-stage renal disease (ESRD) by adolescence, frequently accompanied by sensorineural deafness and ocular complications. Milder forms of ATS also exist. We studied 42 patients (19M, 23F) of nine Hellenic families suspected clinically of X-linked ATS who presented with marked phenotypic heterogeneity. We identified mutations in COL4A5 in six families. Two males with nonsense mutation E228X reached ESRD by ages 14 and 18. Frameshift mutation 2946delT followed the same course with early onset renal involvement and deafness. However, two males with the milder missense mutation G624D, reached ESRD after 39 years and one patient showed thin basement membrane nephropathy (TBMN). Another 5/8 affected males with missense mutation P628L also developed ESRD between 30 and 57 years, while three exhibit only mild chronic renal failure (CRF). The data support previous findings that certain mutations are associated with milder phenotypes and confirm that mutation G624D may be expressed as TBMN with familial hematuria. Similar conclusions apply for missense mutation P628L. Interestingly, mutations G624D and P628L are near the 12th natural interruption of COL4A5 triple helical domain, which may explain the milder phenotype. [ABSTRACT FROM AUTHOR]

Medullary cystic kidney disease type 1 (MCKD1) is an autosomal dominant, tubulo-interstitial nephropathy that causes renal salt wasting and end-stage renal failure in the fourth to seventh decade of life. MCKD1 was localized to chromosome 1q21. We demonstrated haplotype sharing and confirmed the telomeric border by a recombination of D1S2624 in a Belgian kindred. Since the causative gene has been elusive, high resolution haplotype analysis was performed in 16 kindreds. Clinical data and blood samples of 257 individuals (including 75 affected individuals) from 26 different kindreds were collected. Within the defined critical region mutational analysis of 37 genes (374 exons) in 23 MCKD1 patients was performed. In addition, for nine kindreds RT-PCR analysis for the sequenced genes was done to screen for mutations activating cryptic splice sites. We found consistency with the haplotype sharing hypothesis in an additional nine kindreds, detecting three different haplotype subsets shared within a region of 1.19 Mb. Mutational analysis of all 37 positional candidate genes revealed sequence variations in 3 different genes, AK000210, CCT3, and SCAMP3, that were segregating in each affected kindred and were not found in 96 healthy individuals, indicating, that a single responsible gene causing MCKD1 remains elusive. This may point to involvement of different genes within the MCKD1 critical region. [ABSTRACT FROM AUTHOR]

The spectrum of distal renal tubular acidosis (dRTA) includes a genetically heterogeneous group of inherited conditions of both autosomal-dominant and recessive mode of inheritance. The basic defect is linked to the renal part of acid–base homeostasis, which is partly achieved by the regulated luminal secretion of H+ at the apical surface of the α-intercalated cells of renal collecting ducts. This is coupled to bicarbonate reabsorption with chloride counter transport across the basolateral membranes Here, we describe the molecular findings of the first two Greek Cypriot families with recessive dRTA and the long-term clinical findings in four of five affected members. DNA linkage analysis with four polymorphic markers flanking the ATP6V1B1 gene on chromosome 2 gave evidence for positive linkage; direct DNA analysis by automated DNA sequencing revealed that patients in one family were homozygous for mutation 229+1G>T (IVS7+1G>T) and that patients in the second family were compound heterozygous for 229+1G>T and R157C. The mutations were found on four different haplotypes. Both the mutations were previously reported in patients of Turkish origin. Three known polymorphic variants were also identified. The five patients demonstrated the whole clinical spectrum of the disease including death in infancy, failure to thrive, rickets, nephrocalcinosis, nephrolithiasis, and episodes of hypokalemic paralysis. Some of the family members are now in their mid 30s and late 20s, and nephrolithiasis with recurrent renal colics is their main problem. Renal function has remained normal. In conclusion, early diagnosis in infancy and prompt treatment with alkali and potassium supplements is of great benefit to the patient with dRTA and ensures normal growth. The identification of responsible mutations facilitates antenatal or postnatal diagnosis in concerned families and improves the prognosis. [ABSTRACT FROM AUTHOR]

Background. The branchio‐oto‐renal (BOR) syndrome is an autosomal dominant disease characterized by hearing loss of early onset, preauricular pits, branchial clefts, and early progressive chronic renal failure in up to 40% of family affected members. So far, it has not received due attention in the adult European nephrology literature and because of the combination of deafness with chronic renal failure it may be confused with the Alport syndrome. The BOR syndrome is caused by mutations in the EYA1 gene that maps on chromosome 8q13.3. [ABSTRACT FROM PUBLISHER]

The autosomal dominant form of polycystic kidney disease is a very frequent genetically heterogeneous inherited condition affecting approximately 1:1000 individuals of the Caucasian population. The main symptom is the formation of fluid-filled cysts in the kidneys, which grow progressively in size and number with age, and leading to end-stage renal failure in approximately 50% of patients by age 60. About 85% of cases are caused by mutations in the PKD1 gene on chromosome 16p13.3, which encodes for polycystin-1, a membranous glycoprotein with 4302 amino acids and multiple domains. Mutation detection is still a challenge owing to various sequence characteristics that prevent easy PCR amplification and sequencing. Here we attempted a systematic screening of part of the duplicated region of the gene in a large cohort of 53 Hellenic families with the use of single-strand conformation polymorphism analysis of exons 16-34. Our analysis revealed eight most probably disease causing mutations, five deletions and three single amino acid substitutions, in the REJ domain of the protein. In one family, a 3-bp and an 8-bp deletion in exons 20 and 21 respectively, were co-inherited on the same PKD1 chromosome, causing disease in the mother and three sons. Interestingly we did not find any termination codon defects, so common in the unique part of the PKD1 gene. In the same cohort we identified 11 polymorphic sequence variants, four of which resulted in amino acid variations. This supports the notion that the PKD1 gene may be prone to mutagenesis, justifying the relatively high prevalence of polycystic kidney disease. [ABSTRACT FROM AUTHOR]

Mutations in three different genes, PKD1, PKD2 and PKD3, can cause a very similar clinical picture of the autosomal dominant form of polycystic kidney disease (ADPKD). Apparently, mutations in the PKD3 gene, which is still unmapped, are very rare, whereas PKD1 defects account for about 85% of cases. Although ADPKD is a frequent monogenic disorder affecting approximately 1:1000 individuals in the Caucasian population, progress in understanding its pathology was somewhat slow until relatively recently when the PKD1 and PKD2 genes were mapped and cloned. They are both large, being approximately 52 kb and 68 kb in length respectively, and in addition, PKD1 is fairly complex, thus complicating mutation detection. The gene products, polycystin-1 and polycystin-2, are trans-membranous glycoproteins and are considered to be involved in signalling pathways, in cooperation with additional partners. Immunostaining studies in both humans and mice have revealed information regarding the localization of polycystins and their role in the development and maintenance of nephrons. Recent experimentation from various laboratories has shown that loss of heterozygosity and acquired somatic second hits may account, at least partly, for the inter- and intrafamilial phenotypic heterogeneity of the disease, while at the same time, the existence of other modifying loci is also hypothesized. The two-hit hypothesis is admittedly a very attractive one in that it can explain many of the features of the disease, whereas recent data regarding a trans-heterozygous model for cystogenesis adds to the complexity of the molecular mechanisms that can lead to pathogenesis. [ABSTRACT FROM AUTHOR]

Several hereditary disorders, particularly those affecting the physiological anticoagulation systems, have been well established as risk factors for venous thromboembolism. In the present study, we investigated the prevalence of the following thrombogenic mutations in a Greek-Cypriots population: the G1691 factor V Leiden mutation, the G20210A mutation in the prothrombin gene, and the C677T mutation in the methylenetetrahydrofolate reductase (MTHFR). All three variants have been documented to be significant risk factors for various cardiovascular conditions. Ninety unrelated subjects were screened. For the Leiden mutation, 11 subjects (12.2%) were heterozygous and one (1.1%) was homozygous. Seven subjects (7.8%) were heterozygous for the G20210A variant in prothrombin; no homozygotes were identified. The C677T mutation in MTHFR was found in 40 individuals in the heterozygous state (44.4%), and in 16 individuals in the homozygous state (17.8%). These data demonstrate that Greek-Cypriots have an increased frequency of thrombogenic mutations, and suggest that screening for these mutations should be seriously considered, especially when surgery or pregnancy is planned. This is the first study for the frequency of mutations in risk factors that predispose to thrombophilia on the island of Cyprus. [ABSTRACT FROM AUTHOR]

Background. Ultrasound, genetic and clinical correlations are available for ADPKD-1, but lacking for ADPKD-2. The present study was carried out to address: (i) the age-related diagnostic usefulness of ultrasound compared with genetic linkage studies; (ii) the age-related incidence and prevalence of relevant symptoms and complications; and (iii) the age and causes of death in patients with ADPKD-2. [ABSTRACT FROM PUBLISHER]

Loss of heterozygosity (LOH) is a molecular phenomenon that denotes the loss of one of the two alleles at a specific locus. It is frequently associated with tumour suppressor genes in various cancers and also with hyperproliferative disorders, although not exclusively. Interestingly, in conditions where there is an inherited germline mutation, the lost allele is always the functional one, thereby rendering a phenotypically dominant disease of recessive character at the cellular level. A disease more recently shown to be associated with LOH is polycystic kidney disease type 1, a systemic disorder characterized by significant pleiotropy. The main pathology is from renal cyst formation that eventually leads to end-stage renal failure during adult life. We describe the identification of a missense mutation in the repeated part of the PKD1 gene, exon 31, that substitutes valine for methionine. The mutation, M3375V, cosegregates with the disease phenotype in a large Cypriot family. During transplantation of one patient, one of the polycystic kidneys was removed and DNA was isolated from cystic epithelial cells. In 3 of 17 cysts examined with intragenic and flanking polymorphic markers on chromosome 16 we detected LOH, since the wild-type allele was lost, thereby rendering the affected kidneys of mosaic character. The degree of LOH was extensive and varied among the three cysts, supporting the multiplicity of expression of the phenomenon on different occasions. No LOH was detected for other selected loci examined. Our work further supports the hypothesis that the rate-limiting step in cyst formation may be the occurrence of a second somatic hit, although other factors may be also involved. The high frequency of mutations at this locus may, to a great extent, explain the variability in phenotype observed among patients in the same families, and the relatively high frequency of the disease worldwide. [ABSTRACT FROM AUTHOR]

Autosomal dominant polycystic kidney disease (ADPKD) is caused by mutations in one of three genes: PKD1 on chromosome 16 accounts for ∼85% of cases whereas PKD2 on chromosome 4 accounts for ∼15%. Mutations in the PKD3 gene are rare. All patients present with similar clinical phenotypes, and the cardinal symptom is the formation of fluid-filled cysts in the kidneys. Previous work has provided data supporting the notion that cysts in ADPKD1 are focal in nature and form after loss of function of polycystin 1. This became evident by demonstrating that the normal PKD1 allele was inactivated somatically by loss of heterozygosity or by mutagenesis in a subset of renal or liver cysts examined. We show in this report, for the first time, multiple novel somatic mutations within the PKD2 gene of epithelial cells, in both kidneys of an ADPKD2 patient. From a total of 21 cysts examined, seven (33%) had the same C insertion within the inherited wild-type allele. In two other cysts, a nonsense mutation and a splice site AG deletion had occurred in a PKD2 allele that could not be identified as the inherited wild-type or mutant. We suggest that the autosomal dominant form of ADPKD2 occurs by a cellular recessive mechanism, supporting a two-hit model for cyst formation. [ABSTRACT FROM AUTHOR]

The PKD1 gene, which is responsible for the most common form of autosomal dominant polycystic kidney disease, has recently been cloned and sequenced. Many disease-causing mutations have been characterized in this gene, most of them resulting in premature protein termination. However, mutation analysis is not routinely implemented for family investigations in a clinical setting, because of the large size and complexity of the gene. Instead, genetic linkage analysis using highly polymorphic CA dinucleotide repeats that map around the gene is still the method of choice. Recently, a few intragenic polymorphisms have been described that are also useful for linkage studies. Here, a new diallelic polymorphism is described for amino acid residue 4058, Ala/Val4058, with allelic frequencies of 0.88 and 0.12, respectively, and a heterozygosity of 0.23, in the Greek and Greek-Cypriot populations. Interestingly, this polymorphism and Ala4091-A/G, which has previously been described in Caucasians, were not detected in DNA from 44 Japanese samples tested. This is particularly important when allelic frequencies in a particular population are used for linkage analysis of families of different ethnic origin. Also, observation of the two polymorphisms together as haplotypes suggests that the Ala/Val4058 polymorphism occurred more recently than the establishment of the Ala4091-A/G polymorphism, and specifically on the G allele. [ABSTRACT FROM AUTHOR]

Mutations in the PKD1 gene on the short arm of chromosome 16 account for 85%-90% of polycystic kidney disease patients in the Caucasian population. After the recent characterization of the gene, we started a search for mutations in its 3′-end unique portion in Cypriot patients, by using the method of single-strand conformation polymorphism (SSCP). In one large family, we identified a nucleotide substitution at position 12 258 of the cDNA; this substitutes cysteine-4086 by a premature termination codon (C4086X). It has been inherited by every affected family member but not by unaffected members, nor by patients from 13 other Cypriot families. A new polymerase chain reaction (PCR) primer has been designed to engineer a novel DdeI recognition site upon PCR amplification, thereby allowing easy detection of the mutation by PCR-restriction digestion. The premature STOP codon is expected to remove 217 residues from the putative C-terminal intracellular domain of the gene product, polycystin and thus identifies this part as being critical to the production of the disease phenotype, possibly by interfering with the transmission of signals from the extracellular matrix to the cytoplasm. We also describe the identification of the first polymorphism within the encoding region of the gene. It is at alanine 4091, which is encoded by either GCA or GCG. With a heterozygosity of 35%, it should be extremely useful in informative families, especially because the gene lies in an unstable region and is prone to rearrangements. This polymorphism is readily detectable by PCR-restriction digestion with Bsp1286I. [ABSTRACT FROM AUTHOR]

Presents two large Cypriot families that segregate autosomal dominant medullary cystic kidney disease (ADMCKD) with hyperuricemia and gout and with very late age of onset. Characteristics of the disease; Establishment of linkage; Genetic mapping of the region and haplotype analysis; Identification of critical recombinants; Genotyping and DNA mapping.

Polycystic kidney disease is an inherited heterogeneous disorder that affects approximately 1∶1000 Europeans. It is characterized mainly by the formation of cysts in the kidney that lead to end-stage renal failure with late age of onset. Three loci have been identified, PKD1 on the short arm of chromosome 16, which has recently been isolated and characterized, PKD2 on the long arm of chromosome 4, and a third locus of unknown location, that is apparently much rarer. In families that transmit the PKD2 gene there is a significantly later age of onset of symptoms, compared with families that transmit the PKD1 gene, and in general they present with milder progression of symptomatology. For the first time we attempted molecular genetic analysis in seven Cypriot families using highly polymorphic markers around the PKD1 and PKD2 genes. Our data showed that there is genetic and phenotypic heterogeneity among these families. For four of the families we obtained strong evidence for linkage to the PKD1 locus. In two of these families linkage to PKD1 was strengthened by excluding linkage to PKD2 with the use of marker D4S423. In three other families we showed linkage to the PKD2 locus. In the largest of these families one recombinant placed marker D4S1534 distal to D4S231, thereby rendering it the closest proximal marker known to us to date. The application of molecular methods allowed us to make presymptomatic diagnosis for a number of at-risk individuals. [ABSTRACT FROM AUTHOR]

Molecular investigations on a young patient and her family were undertaken to identify the molecular defect responsible for a mild form of osteogenesis imperfecta (OI) with blue sclerae, dentinogenesis imperfecta and joint laxity. Analysis of collagenous proteins from the proband's fibroblasts showed the presence of two populations of α2(I) chains, one normal and one migrating faster on SDS gels, thereby suggesting deletion of amino acid sequences. The faster migrating chains were retained mainly in the cell layer and not found in the extracellular matrix deposited by cultured fibroblasts. Chemical cleavage of mismatch (CCM) analysis on the patient's proα2(I) mRNA:normal cDNA heteroduplexes localized the molecular defect. cDNA sequencing revealed a deletion of exon 20 (54 bp) in about half of the molecules. Genomic DNA sequencing revealed heterozygosity for a G-to-C transversion of the last nucleotide of intron 19, which changed the 3′ consensus splicing site. As a consequence proα2(I)mRNA was abnormally spliced from the last codon of exon 19 to the first codon of exon 21. To our knowledge, this is the first acceptor site mutation so far described in an OI patient. Restriction analysis indicated that the mutation was present also in three other affected family members. The full sequence of COL1A2 introns 19 and 20 are reported. [ABSTRACT FROM AUTHOR]

Cyprus is an island in the eastern Mediter-ranean basin inhabited by people of Caucasian extraction, mostly Greek-Cypriots. The most common inherited disease among Caucasians is cystic fibrosis (CF). Although no careful scientific study had ever been done the impression was that CF was extremely rare among the Greek-Cypriots, with an incidence estimated at around 1∶30,000. About 2 years ago, we introduced molecular diagnostic methodology in an effort to assist clinicians in safer diagnosis of patients presenting with atypical CF symptomatology, and also for testing the hypothesis that mutations that cause milder phenotypes might be responsible for misdiagnosis or for missing entirely some cases of CF. Initial screening for ΔF508 revealed that it is indeed rare in the general population. Further screening of suspected CF patients revealed a novel mutation that converted leucine at position 346 to proline (L346P) in two unrelated families. The second CF mutation was ΔF508 and 1677delTA in the two families respectively, both reportedly associated with severe phenotypes. Yet our patients did not present with typical CF pictures possibly because of the dominant nature of this novel mild mutation in exon 7. Symptoms included failure to thrive, chest infections and electrolyte disturbances. These findings raise the possibility that Cyprus might have been spared very severe CF phenotypes but not cystic fibrosis transmembrane conductance regulator (CFTR) mutations. [ABSTRACT FROM AUTHOR]

A letter to the editor is presented regarding the correlation of genotype-phenotype in the missense mutation in patients with Alport syndrome (AS).

A letter to the editor is presented on cystic fibrosis and the genotypic/phenotypic features in Greek-Cypriots, with emphasis on dehydration as a symptom.

Objective: To search for pyrin mutations associated with familial Mediterranean fever (FMF) in Greece.Patients and Methods: 62 patients fulfilling the Tel Hashomer diagnostic criteria for definite (33) or probable (29) FMF diagnosis were studied. Eight point mutations of pyrin gene were tested by standard methods. Of the 62 patients tested, 48 were Greek, four were Jewish, seven were Armenian, and three were Arab.Results: 42 patients were found to be homozygotes for pyrin mutations; 11 patients were found to carry only one of the tested mutations; in nine patients no mutations were detected.Conclusion: Molecular detection of pyrin gene mutations seems useful in confirming suspected cases, and in detecting asymptomatic cases, of Mediterranean fever in Greece. It may also be used as a screening tool within affected families. [ABSTRACT FROM AUTHOR]

Focuses on the application of molecular genetics to the study of old diseases. Frequency of gene mutations; Differential diagnosis.

Background: Autosomal Dominant Polycystic Kidney Disease (ADPKD) is characterized by the formation of multiple fluid-filled cysts that destroy the kidney architecture resulting in end-stage renal failure. Mutations in genes PKD1 and PKD2 account for nearly all cases of ADPKD. Increased cell proliferation is one of the key features of the disease. Several studies indicated that polycystin-1 regulates cellular proliferation through various signaling pathways, but little is known about the role played by polycystin-2, the product of PKD2. Recently, it was reported that as with polycystin-1, polycystin-2 can act as a negative regulator of cell growth by modulating the levels of the cyclin-dependent kinase inhibitor, p21 and the activity of the cyclin-dependent kinase 2, Cdk2.Methods: Here we utilized different kidney cell-lines expressing wild-type and mutant PKD2 as well as primary tubular epithelial cells isolated from a PKD transgenic rat to further explore the contribution of the p21/Cdk2 pathway in ADPKD proliferation.Results: Surprisingly, over-expression of wild-type PKD2 in renal cell lines failed to inactivate Cdk2 and consequently had no effect on cell proliferation. On the other hand, expression of mutated PKD2 augmented proliferation only in the primary tubular epithelial cells of a rat model but this was independent of the STAT-1/p21 pathway. On the contrary, multiple approaches revealed unequivocally that expression of the cyclin-dependent kinase inhibitor, p57KIP2, is downregulated, while p21 remains unchanged. This p57 reduction is accompanied by an increase in Cdk2 levels.Conclusion: Our results indicate the probable involvement of p57KIP2 on epithelial cell proliferation in ADPKD implicating a new mechanism for mutant polycystin-2 induced proliferation. Most importantly, contrary to previous studies, abnormal proliferation in cells expressing mutant polycystin-2 appears to be independent of STAT-1/p21. [ABSTRACT FROM AUTHOR]

An abstract of the article "Autosomal dominant medullary cystic kidney disease: evidence of gene locus heterogeneity," by A. Fuchshuber, C. C. Deltas, S. Berthold, C. Stavrou, M. Vollmer, C. Burton, T. Feest, D. Krieter, A. Gal, M. Brandis, A. Pierides, and F. Hildebrandt is presented