A novel splicing mutation in the ALB gene causing analbuminaemia in a Portuguese woman

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Diagnosis, Phenotype, and Molecular Genetics of Congenital Analbuminemia

Congenital analbuminemia (CAA) is an inherited, autosomal recessive disorder with an incidence of 1:1,000,000 live birth. Affected individuals have a strongly decreased concentration, or complete absence, of serum albumin. The trait is usually detected by serum protein electrophoresis and immunochemistry techniques. However, due to the existence of other conditions in which the albumin concentrations are very low or null, analysis of the albumin (ALB) gene is necessary for the molecular diagnosis. CAA can lead to serious consequences in the prenatal period, because it can cause miscarriages and preterm birth, which often is due to oligohydramnios and placental abnormalities. Neonatally and in early childhood the trait is a risk factor that can lead to death, mainly from fluid retention and infections in the lower respiratory tract. By contrast, CAA is better tolerated in adulthood. Clinically, in addition to the low level of albumin, the patients almost always have hyperlipidemia, but they usually also have mild oedema, reduced blood pressure and fatigue. The fairly mild symptoms in adulthood are due to compensatory increment of other plasma proteins. The condition is rare; clinically, only about 90 cases have been detected worldwide. Among these, 53 have been studied by sequence analysis of the ALB gene, allowing the identification of 27 different loss of function (LoF) pathogenic variants. These include a variant in the start codon, frame-shift/insertions, frame-shift/deletions, nonsense variants, and variants affecting splicing. Most are unique, peculiar for each affected family, but one, a frame-shift deletion called Kayseri, has been found to cause about one third of the known cases allowing to presume a founder effect. This review provides an overview of the literature about CAA, about supportive and additional physiological and pharmacological information obtained from albumin-deficient mouse and rat models and a complete and up-to-date dataset of the pathogenic variants identified in the ALB gene

Autosomal Dominant Tubulointerstitial Kidney Disease with Adult Onset due to a Novel Renin Mutation Mapping in the Mature Protein

Autosomal dominant tubulointerstitial kidney disease (ADTKD) is a genetically heterogeneous renal disorder leading to progressive loss of renal function. ADTKD-REN is due to rare mutations in renin, all localized in the protein leader peptide and affecting its co-translational insertion in the endoplasmic reticulum (ER). Through exome sequencing in an adult-onset ADTKD family we identified a new renin variant, p.L381P, mapping in the mature protein. To assess its pathogenicity, we combined genetic data, computational and predictive analysis and functional studies. The L381P substitution affects an evolutionary conserved residue, co-segregates with renal disease, is not found in population databases and is predicted to be deleterious by in silico tools and by structural modelling. Expression of the L381P variant leads to its ER retention and induction of the Unfolded Protein Response in cell models and to defective pronephros development in zebrafish. Our work shows that REN mutations outside of renin leader peptide can cause ADTKD and delineates an adult form of ADTKD-REN, a condition which has usually its onset in childhood. This has implications for the molecular diagnosis and the estimated prevalence of the disease and points at ER homeostasis as a common pathway affected in ADTKD-REN, and possibly more generally in ADTKD

A Retrotransposon Insertion in the 5' Regulatory Domain of Ptf1a Results in Ectopic Gene Expression and Multiple Congenital Defects in Danforth’s Short Tail Mouse

Danforth's short tail mutant (Sd) mouse, first described in 1930, is a classic spontaneous mutant exhibiting defects of the axial skeleton, hindgut, and urogenital system. We used meiotic mapping in 1,497 segregants to localize the mutation to a 42.8-kb intergenic segment on chromosome 2. Resequencing of this region identified an 8.5-kb early retrotransposon (ETn) insertion within the highly conserved regulatory sequences upstream of Pancreas Specific Transcription Factor, 1a (Ptf1a). This mutation resulted in up to tenfold increased expression of Ptf1a as compared to wild-type embryos at E9.5 but no detectable changes in the expression levels of other neighboring genes. At E9.5, Sd mutants exhibit ectopic Ptf1a expression in embryonic progenitors of every organ that will manifest a developmental defect: the notochord, the hindgut, and the mesonephric ducts. Moreover, at E 8.5, Sd mutant mice exhibit ectopic Ptf1a expression in the lateral plate mesoderm, tail bud mesenchyme, and in the notochord, preceding the onset of visible defects such as notochord degeneration. The Sd heterozygote phenotype was not ameliorated by Ptf1a haploinsufficiency, further suggesting that the developmental defects result from ectopic expression of Ptf1a. These data identify disruption of the spatio-temporal pattern of Ptf1a expression as the unifying mechanism underlying the multiple congenital defects in Danforth's short tail mouse. This striking example of an enhancer mutation resulting in profound developmental defects suggests that disruption of conserved regulatory elements may also contribute to human malformation syndromes

Molecular Diagnosis of Analbuminemia: A New Case Caused by a Nonsense Mutation in the Albumin Gene

Analbuminemia is a rare autosomal recessive disorder manifested by the absence, or severe reduction, of circulating serum albumin (ALB). We report here a new case diagnosed in a 45 years old man of Southwestern Asian origin, living in Switzerland, on the basis of his low ALB concentration (0.9 g/L) in the absence of renal or gastrointestinal protein loss, or liver dysfunction. The clinical diagnosis was confirmed by a mutational analysis of the albumin (ALB) gene, carried out by single-strand conformational polymorphism (SSCP), heteroduplex analysis (HA), and DNA sequencing. This screening of the ALB gene revealed that the proband is homozygous for two mutations: the insertion of a T in a stretch of eight Ts spanning positions c.1289 + 23–c.1289 + 30 of intron 10 and a c.802 G > T transversion in exon 7. Whereas the presence of an additional T in the poly-T tract has no direct deleterious effect, the latter nonsense mutation changes the codon GAA for Glu244 to the stop codon TAA, resulting in a premature termination of the polypeptide chain. The putative protein product would have a length of only 243 amino acid residues instead of the normal 585 found in the mature serum albumin, but no evidence for the presence in serum of such a truncated polypeptide chain could be obtained by two dimensional electrophoresis and western blotting analysis

Apolipoprotein E in idiopathic nephrotic syndrome and focal segmental glomerulosclerosis

Apolipoprotein E in idiopathic nephrotic syndrome and focal segmental glomerulosclerosis.BackgroundHyperlipemia characterizes nephrotic syndrome (NS) and contributes to the progression of the underlying nephropathy. The data in the literature support an implication of apolipoprotein E (apoE) in both hyperlipemia and focal segmental glomerulosclerosis (FSGS), a malignant condition associated with NS.MethodsThe apoE genotype was determined in 209 nephrotic patients, who were classified according to age and their response to steroids as resistant children (N = 96) and adults (43), and steroid dependent (33) and steroid responder (37) children. A total of 123 presented the histological features of FSGS. In a subgroup of 28 patients, serum and urinary levels of apoE and renal deposits were evaluated by immunofluorescence.ResultsThe allelic frequencies of the three major haplotypes γ2, γ3, and γ4 were the same in nephrotic patients versus controls, and homozygosity for γ3γ3 was comparably the most frequent genotype (70 vs. 71%) followed by γ3γ4, γ2γ3, γ2γ4, γ4γ4. Serum levels of apoE were fivefold higher in NS and in FSGS patients than in controls, with a direct correlation with hypercholesterolemia and proteinuria. ApoE genotypes did not influence serum levels. Urinary levels were 1/10,000 of serum with an increment in nephrotic urines. Finally, immunofluorescence demonstrated the absence of apoE in sclerotic glomeruli, while comparably nephrotic patients with membranous nephropathy had an increased glomerular expression of apoE.ConclusionsApoE is dysregulated in NS with a marked increment in serum, which is a part of the complex lipid metabolism. Down-regulation of glomerular apoE instead is a peculiarity of FSGS and may contribute to the pathogenesis of the disease. The normal distribution of apoE genotypes in nephrotic patients with FSGS excludes a pathogenetic role of genetic variants

Congenital analbuminemia caused by a novel aberrant splicing in the albumin gene

Introduction:Congenital analbuminemia is a rare autosomal recessive disorder manifested by the presence of a very low amount of circulating serum albumin. It is an allelic heterogeneous defect, caused by variety of mutations within the albumin gene in homozygous or compound heterozygous state. Herein we report the clinical and molecular characterization of a new case of congenital analbuminemia diagnosed in a female new-born of consanguineous (first degree cousins) parents from Ankara, Turkey, who presented with a low albumin concentration (< 8 g/L) and severe clinical symptoms. Materials and methods: The albumin gene of the index case was screened by single-strand conformation polymorphism, heteroduplex analysis, and direct DNA sequencing. The effect of the splicing mutation was evaluated by examining the cDNA obtained by reverse transcriptase - polymerase chain reaction (RT-PCR) from the albumin mRNA extracted from proband’s leukocytes. Results: DNA sequencing revealed that the proband is homozygous, and both parents are heterozygous, for a novel G>A transition at position c.1652+1, the first base of intron 12, which inactivates the strongly conserved GT dinucleotide at the 5’ splice site consensus sequence of this intron. The splicing defect results in the complete skipping of the preceding exon (exon 12) and in a frame-shift within exon 13 with a premature stop codon after the translation of three mutant amino acid residues. Conclusions: Our results confirm the clinical diagnosis of congenital analbuminemia in the proband and the inheritance of the trait and contribute to shed light on the molecular genetics of analbuminemia

Teaching molecular genetics: chapter 4—positional cloning of genetic disorders

Positional cloning is the approach of choice for the identification of genetic mutations underlying the pathological development of diseases with simple Mendelian inheritance. It consists of different consecutive steps, starting with recruitment of patients and DNA collection, that are critical to the overall process. A genetic analysis of the enrolled patients and their families is performed, based on genetic recombination frequencies generated by meiotic cross-overs and on genome-wide molecular studies, to define a critical DNA region of interest. This analysis culminates in a statistical estimate of the probability that disease features may segregate in the families independently or in association with specific molecular markers located in known regions. In this latter case, a marker can be defined as being linked to the disease manifestations. The genetic markers define an interval that is a function of their recombination frequencies with the disease, in which the disease gene is localised. The identification and characterisation of chromosome abnormalities as translocations, deletions and duplications by classical cytogenetic methods or by the newly developed microarray-based comparative genomic hybridisation (array CGH) technique may define extensions and borders of the genomic regions involved. The step following the definition of a critical genomic region is the identification of candidate genes that is based on the analysis of available databases from genome browsers. Positional cloning culminates in the identification of the causative gene mutation, and the definition of its functional role in the pathogenesis of the disorder, by the use of cell-based or animal-based experiments. More often, positional cloning ends with the generation of mice with homologous mutations reproducing the human clinical phenotype. Altogether, positional cloning has represented a fundamental step in the research on genetic renal disorders, leading to the definition of several disease mechanisms and allowing a proper diagnostic approach to many conditions