Autosomal dominant ApoA4 mutations present as tubulointerstitial kidney disease with medullary amyloidosis.

Sporadic cases of apolipoprotein A-IV medullary amyloidosis have been reported. Here we describe five families found to have autosomal dominant medullary amyloidosis due to two different pathogenic APOA4 variants. A large family with autosomal dominant chronic kidney disease (CKD) and bland urinary sediment underwent whole genome sequencing with identification of a chr11:116692578 G>C (hg19) variant encoding the missense mutation p.L66V of the ApoA4 protein. We identified two other distantly related families from our registry with the same variant and two other distantly related families with a chr11:116693454 C>T (hg19) variant encoding the missense mutation p.D33N. Both mutations are unique to affected families, evolutionarily conserved and predicted to expand the amyloidogenic hotspot in the ApoA4 structure. Clinically affected individuals suffered from CKD with a bland urinary sediment and a mean age for kidney failure of 64.5 years. Genotyping identified 48 genetically affected individuals; 44 individuals had an estimated glomerular filtration rate (eGFR) under 60 ml/min/1.73 m2, including all 25 individuals with kidney failure. Significantly, 11 of 14 genetically unaffected individuals had an eGFR over 60 ml/min/1.73 m2. Fifteen genetically affected individuals presented with higher plasma ApoA4 concentrations. Kidney pathologic specimens from four individuals revealed amyloid deposits limited to the medulla, with the mutated ApoA4 identified by mass-spectrometry as the predominant amyloid constituent in all three available biopsies. Thus, ApoA4 mutations can cause autosomal dominant medullary amyloidosis, with marked amyloid deposition limited to the kidney medulla and presenting with autosomal dominant CKD with a bland urinary sediment. Diagnosis relies on a careful family history, APOA4 sequencing and pathologic studies.

A mutation in the SAA1 promoter causes hereditary amyloid A amyloidosis.

Amyloid A amyloidosis is a serious clinical condition resulting from the systemic deposition of amyloid A originating from serum amyloid A proteins with the kidneys being the most commonly and earliest affected organ. Previously described amyloid A amyloidosis is linked to increased production and deposition of serum amyloid A proteins secondary to inflammatory conditions arising from infectious, metabolic, or genetic causes. Here we describe a family with primary amyloid A amyloidosis due to a chr11:18287683 T>C (human genome version19) mutation in the SAA1 promoter linked to the amyloidogenic SAA1.1 haplotype. This condition leads to a doubling of the basal SAA1 promoter activity and sustained elevation of serum amyloid A levels that segregated in an autosomal dominant pattern in 12 genetically affected and in none of six genetically unaffected relatives, yielding a statistically significant logarithm of odds (LOD) score over 5. Affected individuals developed proteinuria, chronic kidney disease and systemic deposition of amyloid composed specifically of the SAA1.1 isoform. Tocilizumab (a monoclonal antibody against the interleukin-6 receptor) had a beneficial effect when prescribed early in the disease course. Idiopathic forms represent a significant and increasing proportion (15-20%) of all diagnosed cases of amyloid A amyloidosis. Thus, genetic screening of the SAA1 promoter should be pursued in individuals with amyloid A amyloidosis and no systemic inflammation, especially if there is a positive family history.

Plasma Mucin-1 (CA15-3) Levels in Autosomal Dominant Tubulointerstitial Kidney Disease due to MUC1 Mutations.

INTRODUCTION: Patients with ADTKD-MUC1 have one allele producing normal mucin-1 (MUC1) and one allele producing mutant MUC1, which remains intracellular. We hypothesized that ADTKD-MUC1 patients, who have only 1 secretory-competent wild-type MUC1 allele, should exhibit decreased plasma mucin-1 (MUC1) levels. To test this hypothesis, we repurposed the serum CA15-3 assay used to measure MUC1 in breast cancer to measure plasma MUC1 levels in ADTKD-MUC1. METHODS: This cross-sectional study analyzed CA15-3 levels in a reference population of 6,850 individuals, in 85 individuals with ADTKD-MUC1, and in a control population including 135 individuals with ADTKD-UMOD and 114 healthy individuals. RESULTS: Plasma CA15-3 levels (mean ± standard deviation) were 8.6 ± 4.3 U/mL in individuals with ADTKD-MUC1 and 14.6 ± 5.6 U/mL in controls (p < 0.001). While there was a significant difference in mean CA15-3 levels, there was substantial overlap between the 2 groups. Plasma CA15-3 levels were <5 U/mL in 22% of ADTKD-MUC1 patients, in 0/249 controls, and in 1% of the reference population. Plasma CA15-3 levels were >20 U/mL in 1/85 ADTKD-MUC1 patients, in 18% of control individuals, and in 25% of the reference population. Segregation of plasma CA15-3 levels by the rs4072037 genotype did not significantly improve differentiation between affected and unaffected individuals. CA15-3 levels were minimally affected by gender and estimated glomerular filtration rate. DISCUSSION/CONCLUSIONS: Plasma CA15-3 levels in ADTKD-MUC1 patients are approximately 40% lower than levels in healthy individuals, though there is significant overlap between groups. Further investigations need to be performed to see if plasma CA15-3 levels would be useful in diagnosis, prognosis, or assessing response to new therapies in this disorder.

Autosomal-dominant adult neuronal ceroid lipofuscinosis caused by duplication in DNAJC5 initially missed by Sanger and whole-exome sequencing.

Adult-onset neuronal ceroid lipofuscinoses (ANCL, Kufs disease) are rare hereditary neuropsychiatric disorders characterized by intralysosomal accumulation of ceroid in tissues. The ceroid accumulation primarily affects the brain, leading to neuronal loss and progressive neurodegeneration. Although several causative genes have been identified (DNAJC5, CLN6, CTSF, GRN, CLN1, CLN5, ATP13A2), the genetic underpinnings of ANCL in some families remain unknown. Here we report one family with autosomal dominant (AD) Kufs disease caused by a 30 bp in-frame duplication in DNAJC5, encoding the cysteine-string protein alpha (CSPα). This variant leads to a duplication of the central core motif of the cysteine-string domain of CSPα and affects palmitoylation-dependent CSPα sorting in cultured neuronal cells similarly to two previously described CSPα variants, p.(Leu115Arg) and p.(Leu116del). Interestingly, the duplication was not detected initially by standard Sanger sequencing due to a preferential PCR amplification of the shorter wild-type allele and allelic dropout of the mutated DNAJC5 allele. It was also missed by subsequent whole-exome sequencing (WES). Its identification was facilitated by reanalysis of original WES data and modification of the PCR and Sanger sequencing protocols. Independently occurring variants in the genomic sequence of DNAJC5 encoding the cysteine-string domain of CSPα suggest that this region may be more prone to DNA replication errors and that insertions or duplications within this domain should be considered in unsolved ANCL cases.

Chronic tubulointerstitial kidney disease in untreated adenine phosphoribosyl transferase (APRT) deficiency: A case report
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Adenine phosphoribosyltransferase (APRT) deficiency (OMIM #614723) is a rare autosomal recessive defect in the purine salvage pathway that causes excessive production of 2,8-dihydroxyadenine, leading to nephrolithiasis and chronic kidney disease (CKD). This case report describes the natural history of CKD in untreated APRT deficiency. We describe a novel APRT mutation (chr16:88877985 G / C; c.195 C>/G; p.His54Asp) presenting with CKD without nephrolithiasis. The patient initially required dialysis, but kidney function improved with allopurinol. We reviewed APRT deficiency reported in the literature to determine the loss of kidney function in individuals with untreated APRT deficiency and its relationship to nephrolithiasis. We identified 95 individuals in whom kidney function was assessed prior to treatment. There was a bimodal distribution of kidney failure. AKI occurred frequently in childhood due to obstructing nephrolithiasis or crystalline nephropathy and was usually reversible. CKD developed after age 20 in all patients irrespective of nephrolithiasis history, with 36/42 patients > 40 years of age having at least stage 3 CKD, and 24/42 having an eGFR > 10 mL/min/1.73m2 or being on dialysis. There were 13 adults without nephrolithiasis and 50 adults with nephrolithiasis. The mean age of end-stage renal diesease (ESRD) was 50.52 ± 13.9 for those without nephrolithiasis and 43.4 ± 15.8 years for those with nephrolithiasis (p = 0.24). APRT deficiency is associated with slowly progressive CKD that occurs independently of nephrolithiasis. Diagnosis should be considered in all individuals with chronic tubulointerstitial kidney disease, with or without the presence of nephrolithiasis. In our patient, allopurinol 300 mg/day resulted in improvement of kidney function.
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Noninvasive Immunohistochemical Diagnosis and Novel MUC1 Mutations Causing Autosomal Dominant Tubulointerstitial Kidney Disease.

BACKGROUND: Autosomal dominant tubulointerstitial kidney disease caused by mucin-1 gene (MUC1) mutations (ADTKD-MUC1) is characterized by progressive kidney failure. Genetic evaluation for ADTKD-MUC1 specifically tests for a cytosine duplication that creates a unique frameshift protein (MUC1fs). Our goal was to develop immunohistochemical methods to detect the MUC1fs created by the cytosine duplication and, possibly, by other similar frameshift mutations and to identify novel MUC1 mutations in individuals with positive immunohistochemical staining for the MUC1fs protein. METHODS: We performed MUC1fs immunostaining on urinary cell smears and various tissues from ADTKD-MUC1-positive and -negative controls as well as in individuals from 37 ADTKD families that were negative for mutations in known ADTKD genes. We used novel analytic methods to identify MUC1 frameshift mutations. RESULTS: After technique refinement, the sensitivity and specificity for MUC1fs immunostaining of urinary cell smears were 94.2% and 88.6%, respectively. Further genetic testing on 17 families with positive MUC1fs immunostaining revealed six families with five novel MUC1 frameshift mutations that all predict production of the identical MUC1fs protein. CONCLUSIONS: We developed a noninvasive immunohistochemical method to detect MUC1fs that, after further validation, may be useful in the future for diagnostic testing. Production of the MUC1fs protein may be central to the pathogenesis of ADTKD-MUC1.

Validation of CZECANCA (CZEch CAncer paNel for Clinical Application) for targeted NGS-based analysis of hereditary cancer syndromes.

BACKGROUND: Carriers of mutations in hereditary cancer predisposition genes represent a small but clinically important subgroup of oncology patients. The identification of causal germline mutations determines follow-up management, treatment options and genetic counselling in patients' families. Targeted next-generation sequencing-based analyses using cancer-specific panels in high-risk individuals have been rapidly adopted by diagnostic laboratories. While the use of diagnosis-specific panels is straightforward in typical cases, individuals with unusual phenotypes from families with overlapping criteria require multiple panel testing. Moreover, narrow gene panels are limited by our currently incomplete knowledge about possible genetic dispositions. METHODS: We have designed a multi-gene panel called CZECANCA (CZEch CAncer paNel for Clinical Application) for a sequencing analysis of 219 cancer-susceptibility and candidate predisposition genes associated with frequent hereditary cancers. RESULTS: The bioanalytical and bioinformatics pipeline was validated on a set of internal and commercially available DNA controls showing high coverage uniformity, sensitivity, specificity and accuracy. The panel demonstrates a reliable detection of both single nucleotide and copy number variants. Inter-laboratory, intra- and inter-run replicates confirmed the robustness of our approach. CONCLUSION: The objective of CZECANCA is a nationwide consolidation of cancer-predisposition genetic testing across various clinical indications with savings in costs, human labor and turnaround time. Moreover, the unified diagnostics will enable the integration and analysis of genotypes with associated phenotypes in a national database improving the clinical interpretation of variants.

Multiplex PCR and NGS-based identification of mRNA splicing variants: Analysis of BRCA1 splicing pattern as a model.

Alternative pre-mRNA splicing increases transcriptome plasticity by forming naturally-occurring alternative splicing variants (ASVs). Alterations of splicing processes, caused by DNA mutations, result in aberrant splicing and the formation of aberrant mRNA isoforms. Analyses of hereditary cancer predisposition genes reveal many DNA variants with unknown clinical significance (VUS) that potentially affect pre-mRNA splicing. Therefore, a comprehensive description of ASVs is an essential prerequisite for the interpretation of germline VUS in high-risk individuals. To identify ASVs in a gene of interest, we have proposed an approach based on multiplex PCR (mPCR) amplification of all theoretically possible exon-exon junctions and subsequent characterization of size-selected and pooled mPCR products by next-generation sequencing (NGS). The efficiency of this method is illustrated by a comprehensive analysis of BRCA1 ASVs in human leukocytes, normal mammary, and adipose tissues and stable cell lines. We revealed 94 BRCA1 ASVs, including 29 variants present in all tested samples. While differences in the qualitative expression of BRCA1 ASVs among the analyzed human tissues were minor, larger differences were detected between tissue and cell line samples. Compared with other ASV analysis methods, this approach represents a highly sensitive and rapid alternative for the identification of ASVs in any gene of interest.

Heterozygous Loss-of-Function SEC61A1 Mutations Cause Autosomal-Dominant Tubulo-Interstitial and Glomerulocystic Kidney Disease with Anemia.

Autosomal-dominant tubulo-interstitial kidney disease (ADTKD) encompasses a group of disorders characterized by renal tubular and interstitial abnormalities, leading to slow progressive loss of kidney function requiring dialysis and kidney transplantation. Mutations in UMOD, MUC1, and REN are responsible for many, but not all, cases of ADTKD. We report on two families with ADTKD and congenital anemia accompanied by either intrauterine growth retardation or neutropenia. Ultrasound and kidney biopsy revealed small dysplastic kidneys with cysts and tubular atrophy with secondary glomerular sclerosis, respectively. Exclusion of known ADTKD genes coupled with linkage analysis, whole-exome sequencing, and targeted re-sequencing identified heterozygous missense variants in SEC61A1-c.553A>G (p.Thr185Ala) and c.200T>G (p.Val67Gly)-both affecting functionally important and conserved residues in SEC61. Both transiently expressed SEC6A1A variants are delocalized to the Golgi, a finding confirmed in a renal biopsy from an affected individual. Suppression or CRISPR-mediated deletions of sec61al2 in zebrafish embryos induced convolution defects of the pronephric tubules but not the pronephric ducts, consistent with the tubular atrophy observed in the affected individuals. Human mRNA encoding either of the two pathogenic alleles failed to rescue this phenotype as opposed to a complete rescue by human wild-type mRNA. Taken together, these findings provide a mechanism by which mutations in SEC61A1 lead to an autosomal-dominant syndromic form of progressive chronic kidney disease. We highlight protein translocation defects across the endoplasmic reticulum membrane, the principal role of the SEC61 complex, as a contributory pathogenic mechanism for ADTKD.

Acadian variant of Fanconi syndrome is caused by mitochondrial respiratory chain complex I deficiency due to a non-coding mutation in complex I assembly factor NDUFAF6.

The Acadian variant of Fanconi Syndrome refers to a specific condition characterized by generalized proximal tubular dysfunction from birth, slowly progressive chronic kidney disease and pulmonary interstitial fibrosis. This condition occurs only in Acadians, a founder population in Nova Scotia, Canada. The genetic and molecular basis of this disease is unknown. We carried out whole exome and genome sequencing and found that nine affected individuals were homozygous for the ultra-rare non-coding variant chr8:96046914 T > C; rs575462405, whereas 13 healthy siblings were either heterozygotes or lacked the mutant allele. This variant is located in intron 2 of NDUFAF6 (NM_152416.3; c.298-768 T > C), 37 base pairs upstream from an alternative splicing variant in NDUFAF6 chr8:96046951 A > G; rs74395342 (c.298-731 A > G). NDUFAF6 encodes NADH:ubiquinone oxidoreductase complex assembly factor 6, also known as C8ORF38. We found that rs575462405-either alone or in combination with rs74395342-affects splicing and synthesis of NDUFAF6 isoforms. Affected kidney and lung showed specific loss of the mitochondria-located NDUFAF6 isoform and ultrastructural characteristics of mitochondrial dysfunction. Accordingly, affected tissues had defects in mitochondrial respiration and complex I biogenesis that were corrected with NDUFAF6 cDNA transfection. Our results demonstrate that the Acadian variant of Fanconi Syndrome results from mitochondrial respiratory chain complex I deficiency. This information may be used in the diagnosis and prevention of this disease in individuals and families of Acadian descent and broadens the spectrum of the clinical presentation of mitochondrial diseases, respiratory chain defects and defects of complex I specifically.

Rare variants in known and novel candidate genes predisposing to statin-associated myopathy.

AIM: Genetic variants affecting statin uptake, metabolism or predisposing to muscular diseases may confer susceptibility to statin-induced myopathy. Besides the SLCO1B1 rs4149056 genotype, common genetic variants do not seem to determine statin-associated myopathy. Here we aimed to address the potential role of rare variants. METHODS: We performed whole exome sequencing in 88 individuals suffering from statin-associated myopathy and assessed the burden of rare variants using candidate-gene and exome-wide association analysis. RESULTS: In the novel candidate gene CLCN1, we identified a heterozygote truncating mutation p.R894* in four patients. In addition, we detected predictably pathogenic case-specific variants in MYOT, CYP3A5, SH3TC2, FBXO32 and RBM20. CONCLUSION: These findings support the role of rare variants and nominate loci for follow-up studies.

Variable clinical presentation of an MUC1 mutation causing medullary cystic kidney disease type 1.

BACKGROUND AND OBJECTIVES: The genetic cause of medullary cystic kidney disease type 1 was recently identified as a cytosine insertion in the variable number of tandem repeat region of MUC1 encoding mucoprotein-1 (MUC1), a protein that is present in skin, breast, and lung tissue, the gastrointestinal tract, and the distal tubules of the kidney. The purpose of this investigation was to analyze the clinical characteristics of families and individuals with this mutation. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: Families with autosomal dominant interstitial kidney disease were referred for genetic analysis over a 14-year period. Families without UMOD or REN mutations prospectively underwent genotyping for the presence of the MUC1 mutation. Clinical characteristics were retrospectively evaluated in individuals with the MUC1 mutation and historically affected individuals (persons who were both related to genetically affected individuals in such a way that ensured that they could be genetically affected and had a history of CKD stage IV or kidney failure resulting in death, dialysis, or transplantation). RESULTS: Twenty-four families were identified with the MUC1 mutation. Of 186 family members undergoing MUC1 mutational analysis, the mutation was identified in 95 individuals, 91 individuals did not have the mutation, and111 individuals were identified as historically affected. Individuals with the MUC1 mutation suffered from chronic kidney failure with a widely variable age of onset of end stage kidney disease ranging from 16 to >80 years. Urinalyses revealed minimal protein and no blood. Ultrasounds of 35 individuals showed no medullary cysts. There were no clinical manifestations of the MUC1 mutation detected in the breasts, skin, respiratory system, or gastrointestinal tract. CONCLUSION: MUC1 mutation results in progressive chronic kidney failure with a bland urinary sediment. The age of onset of end stage kidney disease is highly variable, suggesting that gene-gene or gene-environment interactions contribute to phenotypic variability.

Isolated X-linked hypertrophic cardiomyopathy caused by a novel mutation of the four-and-a-half LIM domain 1 gene.

BACKGROUND: Hypertrophic cardiomyopathy with severe left ventricular diastolic dysfunction has been associated with marked exercise intolerance and poor prognosis. However, molecular pathogenesis of this phenotype remains unexplained in a large proportion of cases. METHODS AND RESULTS: We performed whole exome sequencing as an initial genetic test in a large Czech family with 3 males affected by nonobstructive hypertrophic cardiomyopathy with severe left ventricular diastolic dysfunction in end-stage disease. A novel frameshift mutation of four-and-a-half LIM domain 1 gene (FHL1) (c.599_600insT; p.F200fs32X) was detected in these individuals. The mutation does not affect transcription, splicing, and stability of FHL1 mRNA and results in production of truncated FHL1 protein, which is contrary to heart tissue homogenate not detectable in frozen tissue sections of myocardial biopsy of affected males. The identified mutation cosegregated also with abnormal ECG and with 1 case of apical hypertrophic cardiomyopathy in heterozygous females. Although skeletal muscle involvement is a common finding in FHL1-related diseases, we could exclude myopathy in all mutation carriers. CONCLUSIONS: We identified a novel FHL1 mutation causing isolated hypertrophic cardiomyopathy with X-chromosomal inheritance.

Mutations in ANTXR1 cause GAPO syndrome.

The genetic cause of GAPO syndrome, a condition characterized by growth retardation, alopecia, pseudoanodontia, and progressive visual impairment, has not previously been identified. We studied four ethnically unrelated affected individuals and identified homozygous nonsense mutations (c.262C>T [p.Arg88*] and c.505C>T [p.Arg169*]) or splicing mutations (c.1435-12A>G [p.Gly479Phefs*119]) in ANTXR1, which encodes anthrax toxin receptor 1. The nonsense mutations predictably trigger nonsense-mediated mRNA decay, resulting in the loss of ANTXR1. The transcript with the splicing mutation theoretically encodes a truncated ANTXR1 containing a neopeptide composed of 118 unique amino acids in its C terminus. GAPO syndrome's major phenotypic features, which include dental abnormalities and the accumulation of extracellular matrix, recapitulate those found in Antxr1-mutant mice and point toward an underlying defect in extracellular-matrix regulation. Thus, we propose that mutations affecting ANTXR1 function are responsible for this disease's characteristic generalized defect in extracellular-matrix homeostasis.

Dominant renin gene mutations associated with early-onset hyperuricemia, anemia, and chronic kidney failure.

Through linkage analysis and candidate gene sequencing, we identified three unrelated families with the autosomal-dominant inheritance of early onset anemia, hypouricosuric hyperuricemia, progressive kidney failure, and mutations resulting either in the deletion (p.Leu16del) or the amino acid exchange (p.Leu16Arg) of a single leucine residue in the signal sequence of renin. Both mutations decrease signal sequence hydrophobicity and are predicted by bioinformatic analyses to damage targeting and cotranslational translocation of preprorenin into the endoplasmic reticulum (ER). Transfection and in vitro studies confirmed that both mutations affect ER translocation and processing of nascent preprorenin, resulting either in reduced (p.Leu16del) or abolished (p.Leu16Arg) prorenin and renin biosynthesis and secretion. Expression of renin and other components of the renin-angiotensin system was decreased accordingly in kidney biopsy specimens from affected individuals. Cells stably expressing the p.Leu16del protein showed activated ER stress, unfolded protein response, and reduced growth rate. It is likely that expression of the mutant proteins has a dominant toxic effect gradually reducing the viability of renin-expressing cells. This alters the intrarenal renin-angiotensin system and the juxtaglomerular apparatus functionality and leads to nephron dropout and progressive kidney failure. Our findings provide insight into the functionality of renin-angiotensin system and stress the importance of renin analysis in families and individuals with early onset hyperuricemia, anemia, and progressive kidney failure.

Mapping of a new candidate locus for uromodulin-associated kidney disease (UAKD) to chromosome 1q41.

BACKGROUND: Autosomal-dominant juvenile hyperuricemia, gouty arthritis, medullary cysts, and progressive renal insufficiency are features associated with familial juvenile hyperuricemic nephropathy (FJHN), medullary cystic kidney disease type 1 (MCKD1) and type 2 (MCKD2). MCKD1 has been mapped to chromosome 1q21. FJHN and MCKD2 have been mapped to chromosome 16p11.2. FJHN and MCKD2 are allelic, result from uromodulin (UMOD) mutations and the term uromodulin-associated kidney disease (UAKD) has been proposed for them. Linkage studies also reveal families that do not show linkage to any of the identified loci. To identify additional UAKD loci, we analyzed one of these families, with features suggestive of FJHN. METHODS: Clinical, biochemical, and immunohistochemical investigations were used for phenotype characterization. Genotyping, linkage and haplotype analyses were employed to identify the candidate disease region. Bioinformatics and sequencing were used for candidate gene selection and analyses. RESULTS: We identified a new candidate UAKD locus on chromosome 1q41, bounded by markers D1S3470 and D1S1644. We analyzed and found no linkage to this region in eight additional families, who did not map to the previously established loci. We noted that affected individuals showed, in addition to the characteristic urate hypoexcretion, significant reductions in urinary excretion of calcium and UMOD. Immunohistochemical analysis showed that low UMOD excretion resulted from its reduced expression, which is a different mechanism to intracellular UMOD accumulation observed in cases with UMOD mutations. CONCLUSION: We have mapped a new candidate UAKD locus and shown that UAKD may be a consequence of various defects affecting uromodulin biology.

Familial juvenile hyperuricaemic nephropathy (FJHN): linkage analysis in 15 families, physical and transcriptional characterisation of the FJHN critical region on chromosome 16p11.2 and the analysis of seven candidate genes.

Familial juvenile hyperuricaemic nephropathy (FJHN) is an autosomal dominant renal disease characterised by juvenile onset of hyperuricaemia, gouty arthritis, and progressive renal failure at an early age. Recent studies in four kindreds showed linkage of a gene for FJHN to the same genomic interval on chromosome 16p11.2, where the gene for the phenotypically similar medullary cystic disease type 2 (MCKD2) has been localised. In this study we performed linkage analysis in additional 15 FJHN families. Linkage of FJHN to 16p11.2 was confirmed in six families, which suggests that, in a large proportion of FJHN kindreds, the disease is likely to be caused by a gene or genes located outside of 16p11.2. Haplotype analysis of the new and previously analysed families provided two non-overlapping critical regions on 16p11.2-FJHN1, delimited by markers D16S499-D16S3036 and FJHN2, delimited by markers D16S412-D16S3116. Considering MCKD2 to be a distinct molecular entity, the analysis suggests that as many as three kidney disease genes may be located in close proximity on 16p11.2. From genomic databases we compiled integrated physical and transcription maps of whole critical genomic region in which 45 known genes and 129 predicted loci have been localised. We selected, analysed and found no pathogenic mutations in seven candidate genes. The linkage and haplotype analysis reported here demonstrates the genetic heterogeneity of FJHN. The report of integrated physical and mostly in-silico predicted transcription maps of the FJHN critical region provides a basis for precise experimental annotation of the current transcript map, which is essential for final identification of the FJHN gene(s).