Mutations in KEOPS-complex genes cause nephrotic syndrome with primary microcephaly.

Galloway-Mowat syndrome (GAMOS) is an autosomal-recessive disease characterized by the combination of early-onset nephrotic syndrome (SRNS) and microcephaly with brain anomalies. Here we identified recessive mutations in OSGEP, TP53RK, TPRKB, and LAGE3, genes encoding the four subunits of the KEOPS complex, in 37 individuals from 32 families with GAMOS. CRISPR-Cas9 knockout in zebrafish and mice recapitulated the human phenotype of primary microcephaly and resulted in early lethality. Knockdown of OSGEP, TP53RK, or TPRKB inhibited cell proliferation, which human mutations did not rescue. Furthermore, knockdown of these genes impaired protein translation, caused endoplasmic reticulum stress, activated DNA-damage-response signaling, and ultimately induced apoptosis. Knockdown of OSGEP or TP53RK induced defects in the actin cytoskeleton and decreased the migration rate of human podocytes, an established intermediate phenotype of SRNS. We thus identified four new monogenic causes of GAMOS, describe a link between KEOPS function and human disease, and delineate potential pathogenic mechanisms.

Pierson syndrome in an adolescent girl with nephrotic range proteinuria but a normal GFR.

BACKGROUND: Pierson syndrome, caused by mutations in the LAMB2 gene, was originally described as a combination of microcoria and congenital nephrotic syndrome, rapidly progressing to end-stage renal failure. CASE-DIAGNOSIS/TREATMENT: We report a minor variant of Pierson syndrome in a teenage girl with severe myopia since early infancy and proteinuria first detected at age 6. At the age of 11 she was found to carry a unique homozygous non-truncating LAMB2 mutation in exon 2: c.T240G (p.S80R). Renal biopsy revealed mild diffuse mesangial sclerosis and residual expression of laminin ß2. Today at age 14, on treatment with angiotensin-converting enzyme inhibitors and angiotensin receptor blockers, she continues to have nephrotic range proteinuria, but a normal glomerular filtration rate. CONCLUSIONS: LAMB2 mutations should be considered in all patients with glomerular proteinuria and abnormal ocular phenotype, irrespective of age and disease severity.

COQ6 mutations in human patients produce nephrotic syndrome with sensorineural deafness.

Steroid-resistant nephrotic syndrome (SRNS) is a frequent cause of end-stage renal failure. Identification of single-gene causes of SRNS has generated some insights into its pathogenesis; however, additional genes and disease mechanisms remain obscure, and SRNS continues to be treatment refractory. Here we have identified 6 different mutations in coenzyme Q10 biosynthesis monooxygenase 6 (COQ6) in 13 individuals from 7 families by homozygosity mapping. Each mutation was linked to early-onset SRNS with sensorineural deafness. The deleterious effects of these human COQ6 mutations were validated by their lack of complementation in coq6-deficient yeast. Furthermore, knockdown of Coq6 in podocyte cell lines and coq6 in zebrafish embryos caused apoptosis that was partially reversed by coenzyme Q10 treatment. In rats, COQ6 was located within cell processes and the Golgi apparatus of renal glomerular podocytes and in stria vascularis cells of the inner ear, consistent with an oto-renal disease phenotype. These data suggest that coenzyme Q10-related forms of SRNS and hearing loss can be molecularly identified and potentially treated.

Mutations in the human laminin beta2 (LAMB2) gene and the associated phenotypic spectrum.

Mutations of LAMB2 typically cause autosomal recessive Pierson syndrome, a disorder characterized by congenital nephrotic syndrome, ocular and neurologic abnormalities, but may occasionally be associated with milder or oligosymptomatic disease variants. LAMB2 encodes the basement membrane protein laminin beta2, which is incorporated in specific heterotrimeric laminin isoforms and has an expression pattern corresponding to the pattern of organ manifestations in Pierson syndrome. Herein we review all previously reported and several novel LAMB2 mutations in relation to the associated phenotype in patients from 39 unrelated families. The majority of disease-causing LAMB2 mutations are truncating, consistent with the hypothesis that loss of laminin beta2 function is the molecular basis of Pierson syndrome. Although truncating mutations are distributed across the entire gene, missense mutations are clearly clustered in the N-terminal LN domain, which is important for intermolecular interactions. There is an association of missense mutations and small in frame deletions with a higher mean age at onset of renal disease and with absence of neurologic abnormalities, thus suggesting that at least some of these may represent hypomorphic alleles. Nevertheless, genotype alone does not appear to explain the full range of clinical variability, and therefore hitherto unidentified modifiers are likely to exist.

Nineteen novel NPHS1 mutations in a worldwide cohort of patients with congenital nephrotic syndrome (CNS).

BACKGROUND: Recessive mutations in the NPHS1 gene encoding nephrin account for approximately 40% of infants with congenital nephrotic syndrome (CNS). CNS is defined as steroid-resistant nephrotic syndrome (SRNS) within the first 90 days of life. Currently, more than 119 different mutations of NPHS1 have been published affecting most exons. METHODS: We here performed mutational analysis of NPHS1 in a worldwide cohort of 67 children from 62 different families with CNS. RESULTS: We found bi-allelic mutations in 36 of the 62 families (58%) confirming in a worldwide cohort that about one-half of CNS is caused by NPHS1 mutations. In 26 families, mutations were homozygous, and in 10, they were compound heterozygous. In an additional nine patients from eight families, only one heterozygous mutation was detected. We detected 37 different mutations. Nineteen of the 37 were novel mutations (approximately 51.4%), including 11 missense mutations, 4 splice-site mutations, 3 nonsense mutations and 1 small deletion. In an additional patient with later manifestation, we discovered two further novel mutations, including the first one affecting a glycosylation site of nephrin. CONCLUSIONS: Our data hereby expand the spectrum of known mutations by 17.6%. Surprisingly, out of the two siblings with the homozygous novel mutation L587R in NPHS1, only one developed nephrotic syndrome before the age of 90 days, while the other one did not manifest until the age of 2 years. Both siblings also unexpectedly experienced an episode of partial remission upon steroid treatment.

Ophthalmological aspects of Pierson syndrome.

PURPOSE: To study the ocular phenotype of Pierson syndrome and to increase awareness among ophthalmologists of the diagnostic features of this condition. DESIGN: Retrospective, observational case series. METHODS: A multicenter study of 17 patients with molecularly confirmed Pierson syndrome. The eye findings were reviewed and compared to pertinent findings from the literature. RESULTS: The most characteristic ocular anomaly was microcoria. A wide range of additional abnormalities were found, including posterior embryotoxon, megalocornea, iris hypoplasia, cataract, abnormal lens shape, posterior lenticonus, persistent fetal vasculature, retinal detachment, variable axial lengths, and glaucoma. There was high interocular and intrafamilial variability. CONCLUSIONS: Loss-of-function mutations in laminin beta2 (LAMB2) cause a broad range of ocular pathology, emphasizing the importance of laminin beta2 in eye development. Patients with Pierson syndrome can initially present with ocular signs alone. In newborns with marked bilateral microcoria, Pierson syndrome should be considered and renal function investigated.

Analysis of genes encoding laminin beta2 and related proteins in patients with Galloway-Mowat syndrome.

Galloway-Mowat syndrome (GMS) is a rare autosomal recessive disorder characterized by early onset nephrotic syndrome and microcephaly with various anomalies of the central nervous system. GMS likely represents a heterogeneous group of disorders with hitherto unknown genetic etiology. The clinical phenotype to some extent overlaps that of Pierson syndrome (PS), which comprises congenital nephrotic syndrome and distinct ocular abnormalities but which may also include neurodevelopmental deficits and microcephaly. PS is caused by mutations of LAMB2, the gene encoding laminin beta2. We hypothesized that GMS might be allelic to PS or be caused by defects in proteins that interact with laminin beta2. In a cohort of 18 patients with GMS or a GMS-like phenotype we therefore analyzed the genes encoding laminin beta2 (LAMB2), laminin alpha5 (LAMA5), alpha3-integrin (ITGA3), beta1-integrin (ITGB1) and alpha-actinin-4 (ACTN4), but we failed to find causative mutations in these genes. We inferred that LAMA5, ITGA3, ITGB1, and ACTN4 are not directly involved in the pathogenesis of GMS. We excluded LAMB2 as a candidate gene for GMS. Further studies are required, including linkage analysis in families with GMS to identify genes underlying this disease.

Variable phenotype of Pierson syndrome.

Pierson syndrome is caused by mutations in the LAMB2 gene, which encodes the laminin beta2 chain, and is clinically characterized by congenital nephrotic syndrome (CNS) and bilateral microcoria. Here, we describe two cases of Pierson syndrome involving atypical phenotypes. Patient 1 presented with congenital microcoria and infantile nephrotic syndrome. Despite persistent nephrotic syndrome, her renal function was maintained normally until she was 6 years old. Genetic analysis revealed two frame-shifting deletions (truncating mutations) in the LAMB2 gene. Patient 2 presented with isolated CNS without ocular involvement. Her renal function deteriorated progressively over several months, and retinal detachment in the right eye developed when she was aged 10 months. LAMB2 analysis revealed a missense mutation in one allele and a frame-shifting deletion in the other allele. Electron microscopy of a renal biopsy revealed irregular lamellation of the glomerular basement membrane (GBM) in both patients. The phenotypes of Pierson syndrome vary widely, and the severity of the renal phenotype is not always parallel to that of the ocular phenotype. The phenotypic variability likely reflects genotype-phenotype correlations, but unknown genetic or environmental modifiers may play an additional role. Ultrastructural changes of the GBM are a useful diagnostic indicator.

A milder variant of Pierson syndrome.

Congenital nephrotic syndrome (CNS) comprises a heterogeneous group of conditions having in common the disruption of normal glomerular permselectivity, and it carries a poor prognosis, with most patients progressing to end-stage renal disease. Recently, mutations in the LAMB2 gene encoding laminin beta2 were described as the cause of Pierson syndrome, which is characterized by CNS and a complex ocular maldevelopment with microcoria as the most prominent clinical features. Most affected children exhibit early onset of chronic renal failure, neurodevelopmental deficits, and blindness. We report on a patient with CNS, high-grade myopia, and minor structural eye anomalies, including remnants of pupillary membranes, but no microcoria. The patient had not developed renal failure by the age of 16 months, and he showed no neurodevelopmental deficits. He was identified to be homozygous for a novel LAMB2 missense mutation. This observation, together with two recent reports on milder variants of Pierson syndrome, corroborates the concept that the clinical expression of Pierson syndrome is more variable than initially described, and that milder phenotypes may be related to hypomorphic LAMB2 alleles.

SOS1 is the second most common Noonan gene but plays no major role in cardio-facio-cutaneous syndrome.

BACKGROUND: Heterozygous gain-of-function mutations in various genes encoding proteins of the Ras-MAPK signalling cascade have been identified as the genetic basis of Noonan syndrome (NS) and cardio-facio-cutaneous syndrome (CFCS). Mutations of SOS1, the gene encoding a guanine nucleotide exchange factor for Ras, have been the most recent discoveries in patients with NS, but this gene has not been studied in patients with CFCS. METHODS AND RESULTS: We investigated SOS1 in a large cohort of patients with disorders of the NS-CFCS spectrum, who had previously tested negative for mutations in PTPN11, KRAS, BRAF, MEK1 and MEK2. Missense mutations of SOS1 were discovered in 28% of patients with NS. In contrast, none of the patients classified as having CFCS was found to carry a pathogenic sequence change in this gene. CONCLUSION: We have confirmed SOS1 as the second major gene for NS. Patients carrying mutations in this gene have a distinctive phenotype with frequent ectodermal anomalies such as keratosis pilaris and curly hair. However, the clinical picture associated with SOS1 mutations is different from that of CFCS. These findings corroborate that, despite being caused by gain-of-function mutations in molecules belonging to the same pathway, NS and CFCS scarcely overlap genotypically.

Neurodevelopmental deficits in Pierson (microcoria-congenital nephrosis) syndrome.

Pierson syndrome is an autosomal recessive disorder comprising congenital nephrotic syndrome with diffuse mesangial sclerosis and distinct eye abnormalities with microcoria reported as the most prominent clinical feature. LAMB2 mutations leading to lack of laminin beta2 were identified as the molecular cause underlying Pierson syndrome. Although LAMB2 is known to be expressed in the neuromuscular system, and defects of the neuromuscular junctions had been found in laminin beta2-deficient mice, no consistent neurological phenotype has been described clinically in murine or human laminin beta2-deficiency before. This is likely due to the early lethality from renal failure. Here we provide a detailed description of neurological manifestations and development in four patients affected by Pierson syndrome, who survived until the age of 1.3-4.8 years owing to renal replacement therapy. Severe muscular hypotonia, psychomotor retardation, and blindness were present in three patients harboring truncating mutations on both LAMB2 alleles. These symptoms were not attributable to complications of chronic renal failure, thus representing a primary feature of the genetic disorder. Alterations in skeletal muscle tissue from one case were compatible with a chronic denervating process. One affected girl, however, exhibited a milder course of renal disease, normal development, and preserved vision, presumably owing to some residual LAMB2 function. Our findings indicate that severe neurodevelopmental deficits have to be considered as part of Pierson syndrome, at least in the presence of biallelic functional null mutations (complete lack of laminin beta2). This is an important issue in the counseling of parents of an affected newborn or infant.

Fraser and Ablepharon macrostomia phenotypes: concurrence in one family and association with mutated FRAS1.

To date, Fraser syndrome (FS) and Ablepharon macrostomia syndrome (AMS) have been considered distinct disorders, but they share strikingly similar patterns of congenital abnormalities, specifically craniofacial anomalies. While recent research has led to the identification of the genes FRAS1 and FREM2 as the cause of FS, the genetic basis of AMS continues to be enigmatic. We report on the concurrence of AMS-like and Fraser phenotypes in a Brazilian family. Both affected sibs were homozygous for a novel splice site mutation in the FRAS1 gene. Extensive studies on mRNA expression indicated that this mutation most likely leads to loss of function as most previously reported FRAS1 mutations associated with FS. We conclude that a phenotype resembling AMS is a rare clinical expression of FS with no obvious genotype-phenotype correlation. However, the molecular basis of "true" AMS which has been reported as a sporadic disorder in all cases but one, and so far with no relation to FS, is probably different and still needs to be further investigated.

A syndrome comprising childhood-onset glomerular kidney disease and ocular abnormalities with progressive loss of vision is caused by mutated LAMB2.

BACKGROUND: Pierson syndrome (OMIM 609049) is a severe congenital oculorenal disorder with early lethality. The condition is caused by mutations in the LAMB2 gene leading to complete loss of function of the gene product laminin beta2, an essential component of the glomerular and other basement membranes. METHODS: We present a non-consanguineous family with seven offspring affected by childhood-onset nephrotic syndrome progressing to end-stage renal failure and ocular abnormalities including cataracts, anterior chamber and iris abnormalities, and progressive blindness due to retinal detachment. The LAMB2 gene was analysed in this family by direct sequencing. RESULTS: The disorder turned out to segregate with compound heterozygosity for two novel LAMB2 mutations, triangle upV79 and Q1728X. Whereas the mutation Q1728X is predicted to confer complete loss of function, triangle upV79 probably represents a hypomorphic allele, thus explaining the substantially milder phenotype in this family. CONCLUSION: This observation demonstrates that the phenotypic spectrum of LAMB2-associated disorders is broader than previously anticipated, and suggests that milder, non-lethal phenotypes may be associated with mutations retaining some residual function.

Identification of Alu elements mediating a partial PMP22 deletion.

Hereditary neuropathy with liability to pressure palsies (HNPP) is most frequently caused by deletion of a 1.4-Mb region in chromosome 17p11.2-12 including the peripheral myelin protein 22 (PMP22) gene. Smaller deletions partially affecting the PMP22 gene are less frequently observed. We identified in a HNPP patient a deletion of the 5' region of PMP22 including non-coding exon 1, coding exons 2 and 3, whereas, exons 4 and 5 were present. PMP22 exon 3- and 4-specific qPCR resulted in a deletion of one exon 3 allele but in the presence of 2 exon 4 alleles. SNP analysis revealed the presence of heterozygosity for PMP22 coding exons 4 and 5. Finally, MLPA specific for the CMT1A region defined this deletion for the entire 5' region of PMP22 (exons 1, 2 and 3). These partial HNPP deletions may be missed by other techniques, e.g., STR marker analysis. Alu elements have been reported to mediate non-allelic recombination events. Bioinformatic analysis revealed 12 Alu elements flanking in close neighbourhood the estimated 40-kb deletion region as candidates for recombination events. PCR primers were designed to identify a breakpoint-spanning product including the respective Alu elements. PCR-driven identification of a junction fragment was successful with AluJo-AluSq and AluYb9-AluSq specific primer pairs comprising the same intronic region of PMP22. Sequence analysis of these breakpoint-overlapping PCR fragments revealed a 29-bp motif including a chi-like sequence (GCTGG) present both in the AluYb9 and the AluSq element. These data confirm that low-copy repeats (LCRs) mediate non-allelic homologous recombinations (NAHR).