Nine novel <Emphasis Type="Italic">COL4A3</Emphasis> and <Emphasis Type="Italic">COL4A4</Emphasis> mutations and polymorphisms identified in inherited membrane diseases

Both thin basement membrane nephropathy (TBMN) and autosomal recessive Alport syndrome result from mutations in the COL4A3 and COL4A4 genes, and this study documents further mutations and polymorphisms in these genes. Thirteen unrelated children with TBMN and five individuals with autosomal recessive Alport syndrome were examined for mutations in the 52 exons of COL4A3 and the 47 coding exons of COL4A4 using single-stranded conformation polymorphism (SSCP) analysis. Amplicons producing different electrophoretic patterns were sequenced, and mutations were defined as variants that changed an amino acid but were not present in 50 non-hematuric normals. Three further novel mutations were identified. These were IVS 22-5 T>A in the COL4A3 gene in a consanguineous family with autosomal recessive Alport syndrome, and R1677C and R1682Q in the COL4A4 gene. In addition, six novel polymorphisms (G455G, I462I, G736G and IVS 38-8 G>A in COL4A3, and L658L and A1577A in COL4A4) were demonstrated. Many different COL4A3 and COL4A4 mutations cause TBMN and autosomal recessive Alport syndrome. The identification of polymorphisms in these genes is particularly important to enable diagnostic laboratories to distinguish mutations from uncommon normal variants.     

Autosomal dominant Alport syndrome linked to the type IV collage &agr;3 and &agr;4 genes (COL4A3 and COL4A4)

Background: Alport syndrome is a hereditary nephritis that may lead to end-stage renal disease (ERSD) in young adult life and is often associated with sensorineural deafness and/or ocular abnormalities. The majority of families are X-linked due to mutations in the COL4A5 gene at X122. Autosomal forms of the disease are also recognized with recessive disease, having been shown to be due to mutations in the COL4A3 and COL4A4 genes on chromosome 2. Familial benign haematuria has also been mapped to this region in some families.Subjects and methods: We describe a large family with autosomal dominant Alport syndrome in which males and females are equally severely affected and one member with a mild sensorineural deafness reached ESRD aged 35 years. Renal biopsy in four affected patients demonstrated characteristic thickened and split glomerular basement membranes on electron-microscopy. Results: Genetic linkage analysis using markers on chromosome 2q demonstrated co-segregation of the disease with the markers D2S351 and D2S401 with a maximum lod score of 3.4 at zero recombination. Linkage to the COL4A4 gene was confirmed using an intragenic COL4A4 polymorphism. Mutation analysis has revealed a missense Leu36Pro mutation in exon 5 of the adjacent COL4A3 gene in the unaffected mother, which may lead to a more severe phenotype in affected family members carrying this mutation. Conclusion: Mutations in the COL4A3 and COL4A4 cause a spectrum of glomerular basement membrane disease ranging from autosomal recessive Alport syndrome to autosomal dominant Alport syndrome and familial benign haematuria.     

A family with X-linked benign familial hematuria

Gene mutations in COL4A5 located on Xq22 are believed to cause X-linked Alport syndrome, whereas mutations in COL4A3 and COL4A4 located on chromosome 2 are associated with autosomal inherited Alport syndrome or benign familial hematuria. A family with benign familial hematuria caused by COL4A5 mutation, implying X-linked transmission, is reported here for the first time. This result suggests that COL4A5 should be added to the list of causative genes for benign familial hematuria, although the mechanism(s) by which the same mutation leads to the distinct phenotypes, i.e. X-linked Alport syndrome or benign familial hematuria, remains unknown.     

COL4A3/COL4A4 mutations: from familial hematuria to autosomal-dominant or recessive Alport syndrome

COL4A3/COL4A4 mutations: From familial hematuria to autosomal-dominant or recessive Alport syndrome. BACKGROUND: Mutations of the type IV collagen COL4A5 gene cause X-linked Alport syndrome (ATS). Mutations of COL4A3 and COL4A4 have been reported both in autosomal-recessive and autosomal-dominant ATS, as well as in benign familial hematuria (BFH). In the latter conditions, however, clinical features are less defined, few mutations have been reported, and other genes and non-genetic factors may be involved. METHODS: We analyzed 36 ATS patients for COL4A3 and COL4A4 mutations by polymerase chain reaction-single strand conformational polymorphism (PCR-SSCP) and direct sequencing. Sporadic patients who had tested negative for COL4A5 mutations were included with typical cases of autosomal recessive ATS to secure a better definition of the phenotype spectrum. RESULTS: We identified seven previously undescribed COL4A3 mutations: in two genetic compounds and three heterozygotes, and one in COL4A4. In agreement with the literature, some of the mutations of compound heterozygotes were associated with microhematuria in healthy heterozygous relatives. The mutations of heterozygous patients are likely dominant, since no change was identified in the second allele even by sequencing, and they are predicted to result in shortened or abnormal chains with a possible dominant-negative effect. In addition, both genes showed rare variants of unclear pathogenicity, and common polymorphisms that are shared in part with other populations. CONCLUSIONS: This study extends the mutation spectrum of COL4A3 and COL4A4 genes, and suggests a possible relationship between production of abnormal COL IV chains and dominant expression of a continuous spectrum of phenotypes, from ATS to BFH.     

Novel heterozygous COL4A3 mutation in a family with late-onset ESRD

Thin basement membrane nephropathy (TBMN) and Alport syndrome (ATS) are genetically heterogeneous conditions characterized by structural abnormalities in the glomerular basement membrane (GBM). TBMN presents with hematuria, minimal proteinuria, and normal renal function. Although TBMN is an autosomal dominant disease (COL4A3 and COL4A4), ATS can be inherited X-linked (COL4A5), autosomal recessive, or autosomal dominant (both COL4A3 and COL4A4). The clinical course of TBMN is usually benign, whereas ATS typically results in end-stage renal disease (ESRD). Nevertheless, there is a broad spectrum of clinical phenotypes caused by mutations in COL4A3 or COL4A4. We report an Italian family who presented with hematuria and mild proteinuria. Mutational analysis showed a novel heterozygous mutation p.G291E in exon 15 of the COL4A3 gene. Many different mutations in COL4A3 and COL4A4 that cause TBMN have already been identified, but most genetic variability in these genes has been found to cause autosomal ATS. A valid genotype–phenotype correlation for TBMN or ATS is not yet known. Therefore, it is important to identify new mutations by direct sequencing to clarify their clinical importance, to assess the prognosis of the disease, and to avoid renal biopsy.     

COL4A3/COL4A4 Mutations and Features in Individuals with Autosomal Recessive Alport Syndrome

Alport syndrome is an inherited disease characterized by hematuria, progressive renal failure, hearing loss, and ocular abnormalities. Autosomal recessive Alport syndrome is suspected in consanguineous families and when female patients develop renal failure. Fifteen percent of patients with Alport syndrome have autosomal recessive inheritance caused by two pathogenic mutations in either COL4A3 or COL4A4. Here, we describe the mutations and clinical features in 40 individuals including 9 children and 21 female individuals (53%) with autosomal recessive inheritance indicated by the detection of two mutations. The median age was 31 years (range, 6–54 years). The median age at end stage renal failure was 22.5 years (range, 10–38 years), but renal function was normal in nine adults (29%). Hearing loss and ocular abnormalities were common (23 of 35 patients [66%] and 10 of 18 patients [56%], respectively). Twenty mutation pairs (50%) affected COL4A3 and 20 pairs affected COL4A4. Of the 68 variants identified, 39 were novel, 12 were homozygous changes, and 9 were present in multiple individuals, including c.2906C>G (p.(Ser969*)) in COL4A4, which was found in 23% of the patients. Thirty-six variants (53%) resulted directly or indirectly in a stop codon, and all 17 individuals with early onset renal failure had at least one such mutation, whereas these mutations were less common in patients with normal renal function or late-onset renal failure. In conclusion, patient phenotypes may vary depending on the underlying mutations, and genetic testing should be considered for the routine diagnosis of autosomal recessive Alport syndrome.Alport syndrome is an inherited renal disease characterized by hematuria, progressive renal failure, hearing loss, and ocular abnormalities. Alport commented in 1927 that the occurrence of hematuria and hearing loss in a pedigree was not coincidental but represented a clinical syndrome, and that the more severe disease in male individuals was consistent with X-linked inheritance.¹ We now understand that nearly 85% of patients have X-linked disease due to a pathogenic mutation in the COL4A5 gene, and the remaining individuals usually have autosomal recessive inheritance with two pathogenic mutations in either the COL4A3 or COL4A4 gene.Alport syndrome is usually suspected when the typical clinical features are present. Diagnostic features² include a positive family history, a lamellated glomerular basement membrane (GBM),³ high tone sensorineural hearing loss, and lenticonus and macular flecks on ophthalmoscopy.⁴ However, these features do not distinguish between X-linked and autosomal inheritance. The possibility of autosomal recessive disease is often overlooked, but its recognition is important because the genetic implications are different for the patient and other family members. Affected male individuals with X-linked disease, but few female individuals, eventually develop renal failure and the disease is transmitted from one generation to another. With autosomal recessive inheritance, male and female individuals are equally likely to be affected; renal failure tends to occur in only one generation except in the presence of multiple consanguinity. In our previous report of 206 patients referred for molecular testing of COL4A5, the pathogenic mutation detection rates in families fulfilling none, one, two, three, or four diagnostic criteria were 0%, 18%, 64%, 89%, and 81%, respectively. Autosomal recessive inheritance was suspected to account for the families meeting four diagnostic criteria in whom no pathogenic COL4A5 mutation was detected.⁵Nearly 300 pathogenic mutations have been described in the COL4A3 and COL4A4 genes (Leiden Open Variation Database; https://grenada.lumc.nl/LOVD2/COL4A/home.php?action=switch_db), but many of these are from patients with thin basement membrane nephropathy (TBMN). There are few reports describing two pathogenic mutations in individuals with autosomal recessive Alport syndrome.^6–16 Even fewer studies have examined how mutations may determine clinical features.Here we describe genetic mutations and clinical features in 40 patients in whom two pathogenic mutations were identified in the COL4A3 or COL4A4 gene, consistent with the diagnosis of autosomal recessive Alport syndrome. In many cases, the mutations were demonstrated to be in trans, which is on different chromosomes, confirming autosomal recessive inheritance. Testing examined the entire coding region and splice sites of both COL4A3 and COL4A4 using unidirectional fluorescent Sanger DNA sequencing, analyzed using Mutation Surveyor software. For detecting point mutations in the regions screened, this approach has an analytical sensitivity and specificity of >99%.¹⁷     

Mutations in theCOL4A4 and COL4A3 genes cause familial benign hematuria

Familial benign hematuria (FBH) is a common autosomal dominant disorder characterized by the presence of persistent or recurrent hematuria. The clinical and pathologic features of this syndrome resemble those of early Alport syndrome (AS), and for this reason a common molecular defect has been proposed. The COL4A3/4 genes seem to be involved in both autosomal AS and FBH. This study involves a linkage analysis for the COL4A3/4 loci and a search for mutations within these genes in 11 biopsy-proven FBH families. Haplotype analysis showed that linkage to the COL4A3/4 locus could not be excluded in eight of nine families. One family was not linked to this locus; however, it included three affected women who could be X-linked AS carriers. Two families were too small to perform linkage analysis. COL4A3 and COL4A4 mutation screening disclosed six new pathogenic mutations, two in the COL4A3 gene (G985V and G1015E) and four in the COL4A4 gene (3222insA, IVS23-1G>C, 31del11, and G960R). It is the first time that mutations within the COL4A3 gene are described in families with FBH. This study clearly demonstrates the main role of the COL4A4 and COL4A3 genes in the pathogenesis of FBH.     

Structure of the human type IV collagen gene COL4A3 and mutations in autosomal Alport syndrome

Mutations in either the COL4A3 or the COL4A4 genes, encoding the alpha3 and alpha4 chains of type IV collagen, are responsible for the autosomal-recessive form of Alport syndrome, a progressive hematuric nephropathy characterized by glomerular basement membrane abnormalities. Reported here are the complete COL4A3 exon-intron structure and a comprehensive screen for mutations of the 52 COL4A3 exons in 41 unrelated patients diagnosed as having autosomal Alport syndrome. This resulted in the identification of 21 mutations that are expected to be causative. Furthermore, it is shown that heterozygous COL4A3 missense mutations, when symptomatic, can be associated with a broad range of phenotypes, from familial benign hematuria to the complete features of Alport syndrome nephropathy.     

Segregation of hematuria in thin basement membrane disease with haplotypes at the loci for Alport syndrome

BACKGROUND: Inherited hematuria is common and is usually attributed to thin basement membrane disease (TBMD). The aim of this study was to determine how often hematuria in families with TBMD segregated with haplotypes at the chromosomal loci for autosomal recessive and X-linked Alport syndrome (COL4A3/COL4A4 and COL4A5, respectively). METHODS: The families of 22 individuals with TBMD on renal biopsy and with urinary glomerular red blood cell (RBC) counts of more than 50,000/mL were studied using phase-contrast microscopy of the urine and DNA microsatellite markers. Eighteen families had at least two members with hematuria. RESULTS: Hematuria segregated with or was consistent with segregation at the COL4A3/COL4A4 locus in eight (36%) families (P < 0.05 in 5 of these) and at the COL4A5 locus in four (18%) families (P < 0.05 in 2). The lack of segregation in the other 10 (45%) families may have occurred because of incomplete penetrance of the hematuria, de novo mutations, coincidental hematuria in other family members, or the presence of a novel gene locus. In four different families, three of which had hematuria that segregated with the COL4A3/COL4A4 locus, four family members with the hematuria haplotype had spouses with coincidental hematuria (4 of 29, 14%). However, none of their four offspring who had also inherited the hematuria haplotype had the clinical features of autosomal recessive Alport syndrome. CONCLUSIONS: Hematuria in families with TBMD commonly segregates with the COL4A3/COL4A4 locus and thus results from mutations in the same genes as autosomal recessive Alport syndrome. Sometimes TBMD may be confused with the carrier state for X-linked Alport syndrome. However, nearly half of the families in this study had hematuria that did not segregate with the loci for either autosomal recessive or X-linked Alport syndrome.     

A novel model of autosomal dominant Alport syndrome in Dalmatian dogs.

BACKGROUND: Autosomal dominant Alport syndrome is a rare inherited disease characterized clinically by haematuria, renal failure and deafness, and ultrastructurally by a lamellated glomerular basement membrane (GBM). It is usually caused by mutations in the COL4A3 or COL4A4 genes which code for the alpha3 and alpha4 chains of type IV collagen. We describe here a novel spontaneous model of autosomal dominant Alport syndrome in Dalmatian dogs. METHODS: Affected dogs were identified by a urinary protein creatinine >/=0.3. A total of 10 affected adult Dalmatians and eight unaffected age- and sex-matched dogs from breeds other than Dalmatians were examined. In addition, kidneys from five Dalmatian fetuses from affected mothers were examined histologically and ultrastructurally. RESULTS: All affected dogs were purebred Dalmatians and had a common progenitor. Successive generations were affected, and males and females were affected equally often and equally severely, consistent with autosomal dominant inheritance. The median age at onset of renal failure was 18 months (range 8 months to 7 years). Affected dogs were not clinically deaf, and did not have the ocular abnormalities seen in human X-linked or autosomal recessive Alport syndrome. In addition, they did not have the leucocyte inclusions, low platelet counts or large platelets seen in autosomal dominant hereditary nephritis due to MYH9 mutations. The renal histology and ultrastructural appearance of the GBM appeared to be normal in utero. However, affected adult kidneys demonstrated segmental glomerular hyalinosis and sclerosis with tubulo-interstitial inflammation and fibrosis, and on ultrastructural examination the GBM was lamellated with subepithelial frilling, vacuolation and occasional intramembranous deposits. All alpha1(IV)-alpha5(IV) type IV collagen chains were present in the affected GBM and Bowman's capsule. CONCLUSIONS: Autosomal dominant Alport syndrome in Dalmatians resembles the disease in Bull terriers but has arisen independently. These models will enable us to determine how genetic mutations affect the corresponding proteins and overall membrane structure in autosomal dominant Alport syndrome.     

Autosomal recessive Alport syndrome: an in-depth clinical and molecular analysis of five families.

BACKGROUND: Alport syndrome (ATS) is a progressive inherited nephropathy characterized by irregular thinning, thickening and splitting of the glomerular basement membrane (GBM) often associated with hearing loss and ocular symptoms. ATS has been shown to be caused by COL4A5 mutations in its X-linked form and by COL4A3 and COL4A4 mutations in its autosomal forms. METHODS: Five families with a suspicion of ATS were investigated both from a clinical and molecular point of view. COL4A3 and COL4A4 genes were analysed by DHPLC. Automated sequencing was performed to identify the underlying mutation. RESULTS: Molecular analysis indicated that in all 5 cases the correct diagnosis was autosomal recessive ATS. In three families in which parental consanguinity clearly pinpointed to autosomal recessive ATS, we found COL4A4 homozygous mutations in two of them and COL4A3 homozygous mutation in the other one. In the remaining two families a differential diagnosis including X-linked ATS, autosomal recessive ATS and thin basement membrane nephropathy was considered. The molecular analysis demonstrated that the probands were genetic compounds for two different mutations in the COL4A4 gene pinpointing to the correct diagnosis of autosomal recessive ATS. CONCLUSIONS: A clinical evaluation of probands and their relatives of the five families carrying mutations in either the COL4A3 or the COL4A4 gene was carried out to underline the natural history of the autosomal recessive ATS. In addition, this paper stresses the complexity of the clinics and genetics of ATS and how a correct diagnosis is based on a combination of: (i) an in-depth clinical investigation; (ii) a detailed formal genetic analysis; (iii) a correct technical choice of the gene to be investigated; (iv) a correct technical choice of the family member to be included in the mutational screening. A correct diagnosis is the basis for an appropriate genetic counselling dealing with both the correct prognosis and the accurate recurrence risk for the patients and family members.     

Autosomal-dominant Alport syndrome: natural history of a disease due to COL4A3 or COL4A4 gene

BACKGROUND: Alport syndrome is a clinically and genetically heterogeneous nephropathy. The majority of cases are transmitted as an X-linked semidominant condition due to COL4A5 mutations. In this form males are more severely affected than females. Less than 10% of cases are autosomal recessive due to mutation in either COL4A3 or COL4A4. In this rarer form, both males and females are severely affected. Only two cases of autosomal-dominant Alport syndrome have been reported, one due to a COL4A3 mutation and the other due to a COL4A4 mutation. Because of the paucity of the reported families, the natural history of autosomal-dominant Alport syndrome is mostly unknown. METHODS: Four families with likely autosomal-dominant Alport syndrome were investigated. COL4A3 and COL4A4 genes were analyzed by denaturing high-performance liquid chromatography (HPLC). Automated sequencing was performed to identify the underlying mutation. RESULTS: Two families had a mutation in the COL4A4 gene and two in the COL4A3. Accurate clinical evaluation of family members showed interesting results. Affected individuals (22 persons) had a wide range of phenotypes from end-stage renal disease (ESRD) in the fifth decade to a nonprogressive isolated microhematuria. Finally, three heterozygous individuals (90, 22 and 11 years old, respectively) were completely asymptomatic. CONCLUSION: This paper demonstrated that patients affected by autosomal-dominant Alport syndrome have a high clinical variability. Moreover, a reduced penetrance of about 90% (3 of 25) may be considered for the assessment of recurrence risk during genetic counseling of these families.     

Mutations in the COL4A4 gene in thin basement membrane disease

BACKGROUND: Patients with thin basement membrane disease (TBMD) are often from families where hematuria segregates with the COL4A3 and COL4A4 genes. These genes also are affected in autosomal recessive Alport syndrome. The aim of this study was to demonstrate COL4A4 mutations in TBMD. METHODS: Forty-eight unrelated individuals with TBMD who had no family members with autosomal recessive Alport syndrome were examined for COL4A4 mutations. The diagnosis of TBMD had been confirmed by renal biopsy (43/48, 90%) or by a family history of hematuria but without a renal biopsy (5/48, 10%). The 47 coding exons of COL4A4 were screened for mutations with the methods of enzyme mismatch cleavage or single stranded conformational polymorphism (SSCP) analysis, and exons that demonstrated electrophoretic abnormalities were sequenced. RESULTS: Nine variants that altered the coding sequences were identified. These were nonsense and frameshift mutations that resulted in stop codons (N = 3), and glycine (N = 3) and non-glycine missense variants (N = 3). Four intronic variants and three neutral polymorphisms were also detected. In total, four variants were considered 'pathogenic' principally because they resulted in stop codons or were not present in non-hematuric normal subjects. Three variants were considered 'possibly pathogenic' but two of these were each present in one of 46 non-hematuric normal subjects. CONCLUSIONS: Pathogenic COL4A4 mutations were demonstrated in three of the nine (33%) families in whom hematuria segregated with the COL4A3/COL4A4 locus. Two stop codons (R1377X and 2788/91delG) and a glycine substitution (G960R) resulted in hematuria in all 16 members who were tested from these three families. The S969X mutation described here in TBMD for the first time, as well as the R1377X mutation, also occur in autosomal recessive Alport syndrome.     

Diagnosis of Alport syndrome without biopsy?

Alport syndrome (AS) is genetically heterogeneous. The gene COL4A5 is mutated in the more frequent X-linked dominant form of the disease whereas COL4A3 or COL4A4 are mutated in the autosomal recessive and dominant forms. Diagnosis of AS and determination of the mode of transmission are important because of the differences in prognosis and genetic counselling attached to these different forms. Recently, promising results have been obtained in Col4a3-null mice, an animal model for AS, with different therapeutic trials when administered early in the course of the disease, an additional reason for making early diagnosis of AS in children. Since the identification of the molecular basis of the disease, mutation screening is theoretically the best diagnostic approach, avoiding the use or renal or skin biopsy. However, for many reasons linked to the genetic heterogeneity of the disease, the large size of the three genes and the random distribution of the mutations all along these huge genes, this method is tedious, expensive and time consuming. Moreover, its sensitivity is reduced. For these reasons, evaluation of the expression of type IV collagen chains in the skin, and if necessary in the renal basement membrane, remains a useful tool for AS diagnosis. At this time, the indication for these different approaches, which are not mutually exclusive but complementary, depends on the patient clinical presentation and family history.     

The role of molecular genetics in diagnosing familial hematuria(s)

Familial microscopic hematuria (MH) of glomerular origin represents a heterogeneous group of monogenic conditions involving several genes, some of which remain unknown. Recent advances have increased our understanding and our ability to use molecular genetics for diagnosing such patients, enabling us to study their clinical characteristics over time. Three collagen IV genes, COL4A3, COL4A4, and COL4A5 explain the autosomal and X-linked forms of Alport syndrome (AS), and a subset of thin basement membrane nephropathy (TBMN). A number of X-linked AS patients follow a milder course reminiscent of that of patients with heterozygous COL4A3/COL4A4 mutations and TBMN, while at the same time a significant subset of patients with TBMN and familial MH progress to chronic kidney disease (CKD) or end-stage kidney disease (ESKD). A mutation in CFHR5, a member of the complement factor H family of genes that regulate complement activation, was recently shown to cause isolated C3 glomerulopathy, presenting with MH in childhood and demonstrating a significant risk for CKD/ESKD after 40?years old. Through these results molecular genetics emerges as a powerful tool for a definite diagnosis when all the above conditions enter the differential diagnosis, while in many at-risk related family members, a molecular diagnosis may obviate the need for another renal biopsy.     

Animal models of Alport syndrome.

Introduction The last decade of the twentieth century was a very productiveperiod for the study of Alport syndrome. Alport syndrome wasshown to result from mutations in certain members of the typeIV collagen family of proteins, the 3(IV), 4(IV), and 5(IV)chains. Several hundred different mutations in the COL4A5 gene,which encodes the 5(IV) chain, were described in patients withX-linked Alport syndrome [1]. A few dozen mutations were foundin the COL4A3 and COL4A4 genes, which respectively encode the3(IV) and 4(IV) chains, in patients with autosomal recessiveAlport syndrome [2,3]. Autosomal dominant Alport syndrome, dueto heterozygous mutations in COL4A3 or COL4A4, was establishedas an entity, and distinguished from Fechtner and Epstein syndromes,which arise from mutations in a non-collagen locus, MYH9 [4–7].Investigators established that mutations in the 3(IV), 4(IV),or      

Glomerular pathology in Alport syndrome: a molecular perspective

We have known for some time that mutations in the genes encoding 3 of the 6 type IV collagen chains are the underlying defect responsible for both X-linked (where the COL4A5 gene is involved) and autosomal (where either COL4A3 or COL4A4 genes are involved) Alport syndrome. The result of these mutations is the absence of the sub-epithelial network of all three chains in the glomerular basement membrane (GBM), resulting, at maturity, in a type IV collagen GBM network comprising only α1(IV) and α2(IV) chains. The altered GBM functions adequately in early life. Eventually, there is onset of proteinuria associated with the classic and progressive irregular thickening, thinning, and splitting of the GBM, which culminates in end-stage renal failure. We have learned much about the molecular events associated with disease onset and progression through the study of animal models for Alport syndrome, and have identified some potential therapeutic approaches that may serve to delay the onset or slow the progression of the disease. This review focuses on where we are in our understanding of the disease, where we need to go to understand the molecular triggers that set the process in motion, and what emergent therapeutic approaches show promise for ameliorating disease progression in the clinic.     

Identification of 47 novel mutations in patients with Alport syndrome and thin basement membrane nephropathy

Background

Alport syndrome (ATS) is a progressive hereditary nephropathy characterized by hematuria and proteinuria. It can be associated with extrarenal manifestations. In contrast, thin basement membrane nephropathy (TBMN) is characterized by microscopic hematuria, is largely asymptomatic, and is rarely associated with proteinuria and end-stage renal disease. Mutations have been identified in the COL4A5 gene in ATS and in the COL4A3 and COL4A4 genes in ATS and TBMN. To date, more than 1000 different mutations in COL4A5, COL4A3, and COL4A4 are known.

Methods

In this study mutational analysis by exon sequencing and multiplex ligation-dependent probe amplification was performed in a large European cohort of families with ATS and TBMN.

Results

Molecular diagnostic testing of 216 individuals led to the detection of 47 novel mutations, thereby expanding the spectrum of known mutations causing ATS and TBMN by up to 10 and 6 %, respectively, depending on the database. Remarkably, a high number of ATS patients with only single mutations in COL4A3 and COL4A4 were identified. Additionally, three ATS patients presented with synonymous sequence variants that possible affect correct mRNA splicing, as suggested by in silico analysis.

Conclusions

The results of this study clearly broaden the genotypic spectrum of known mutations for ATS and TBMN, which will in turn now facilitate future studies into genotype–phenotype correlations. Further studies should also examine the significance of single heterozygous mutations in COL4A3 and COL4A4 and of synonymous sequence variants associated with ATS.