Identification of a novel deletion in the MMAA gene in two Iranian siblings with vitamin B12-responsive methylmalonic acidemia.

BACKGROUND: Adenosylcobalamin (vitamin B12) is a coenzyme required for the activity of methylmalonyl-CoA mutase. Defects in this enzyme are a cause of methylmalonic acidemia (MMA). Methylmalonic acidemia, cblA type, is an inborn error of vitamin B12 metabolism that occurs due to mutations in the MMAA gene. MMAA encodes the enzyme which is involved in translocation of cobalamin into the mitochondria. METHODS: One family with two MMA-affected children, one unaffected child, and their parents were studied. The two affected children were diagnosed by urine organic acid analysis using gas chromatography-mass spectrometry. MMAA was analyzed by PCR and sequencing of its coding region. RESULTS: A homozygous deletion in exon 4 of MMAA, c.674delA, was found in both affected children. This deletion causes a nucleotide frame shift resulting in a change from asparagine to methionine at amino acid 225 (p.N225M) and a truncated protein which loses the ArgK conserved domain site. mRNA expression analysis of MMAA confirmed these results. CONCLUSION: We demonstrate that the deletion in exon 4 of the MMAA gene (c.674 delA) is a pathogenic allele via a nucleotide frame shift resulting in a stop codon and termination of protein synthesis 38 nucleotides (12 amino acids) downstream of the deletion.

Whole Exome Sequencing Reveals a Novel Damaging Mutation in Human Fibroblast Activation Protein in a Family with Esophageal Squamous Cell Carcinoma.

PURPOSE: Esophageal squamous cancer cell (ESCC), with late diagnosis and poor rate of survival, is a significant cause of mortality in the developing countries. The hypothesis of rare high penetrance with mutations in new genes may explain the underlying predisposition in some of these familial cases. METHODS: Exome sequencing was performed in the patients with ESCC with strong disease aggregation, two sisters with ESCC cancer, and one with breast cancer. Data analysis selected only very rare variants (0-0.1%) located in genes with a role compatible with cancer. In addition, the homology modeling of the novel mutation (A459D) discovered in FAP gene was performed by using the online Swiss-Prot server for automated modeling and the resulted structure has been modified and analyzed by using bioinformatics software to thoroughly study the structural deficiencies caused by the novel mutation. RESULTS: Ten final candidate variants were selected and six genes validated by Sanger sequencing. Correct family segregation and somatic studies were used to categorize the most interesting variants in FAP, BOD1L, RAD51, Gasdermin D, LGR5, and CERS4. A novel, human mutation C1367A encoding Ala459 Asp (accession number: KT988039), occurring in the blade of the ß propeller domain, was identified in two sisters with ESCC. CONCLUSIONS: We identified novel mutations in three drug delivery genes, a tumor suppressor and also a stem cell marker of esophageal that may have a role in cancer treatment and are involved in cellular pathways, which supports their putative involvement in germ-line predisposition to this neoplasm.

Four novel mutations of the BCKDHA, BCKDHB and DBT genes in Iranian patients with maple syrup urine disease.

BACKGROUND: Maple syrup urine disease (MSUD) is a rare metabolic autosomal recessive disorder caused by dysfunction of the branched-chain α-ketoacid dehydrogenase (BCKDH) complex. Mutations in the BCKDHA, BCKDHB and DBT genes are responsible for MSUD. The current study analyzed seven Iranian MSUD patients genetically and explored probable correlations between their genotype and phenotype. METHODS: The panel of genes, including BCKDHA, BCKDHB and DBT, was evaluated, using routine the polymerase chain reaction (PCR)-sequencing method. In addition, protein modeling (homology and threading modeling) of the deduced novel mutations was performed. The resulting structures were then analyzed, using state-of-the-art bioinformatics tools to better understand the structural and functional effects caused by mutations. RESULTS: Seven mutations were detected in seven patients, including four novel pathogenic mutations in BCKDHA (c.1198delA, c.629C>T), BCKDHB (c.652C>T) and DBT (c.1150A>G) genes. Molecular modeling of the novel mutations revealed clear changes in the molecular energy levels and stereochemical traits of the modeled proteins, which may be indicative of strong correlations with the functional modifications of the genes. Structural deficiencies were compatible with the observed phenotypes. CONCLUSIONS: Any type of MSUD can show heterogeneous clinical manifestations in different ethnic groups. Comprehensive molecular investigations would be necessary for differential diagnosis.

Novel mutations and their genotype-phenotype correlations in patients with Noonan syndrome, using next-generation sequencing.

PURPOSE: Noonan Syndrome (NS) is an autosomal dominant disorder with many variable and heterogeneous conditions. The genetic basis for 20-30% of cases is still unknown. This study evaluates Iranian Noonan patients both clinically and genetically for the first time. MATERIALS/METHODS: Mutational analysis of PTPN11 gene was performed in 15 Iranian patients, using PCR and Sanger sequencing at phase one. Then, as phase two, Next Generation Sequencing (NGS) in the form of targeted resequencing was utilized for analysis of exons from other related genes. Homology modelling for the novel founded mutations was performed as well. The genotype, phenotype correlation was done according to the molecular findings and clinical features. RESULTS: Previously reported mutation (p.N308D) in some patients and a novel mutation (p.D155N) in one of the patients were identified in phase one. After applying NGS methods, known and new variants were found in four patients in other genes, including: CBL (p. V904I), KRAS (p. L53W), SOS1 (p. I1302V), and SOS1 (p. R552G). Structural studies of two deduced novel mutations in related genes revealed deficiencies in the mutated proteins. Following genotype, phenotype correlation, a new pattern of the presence of intellectual disability in two patients was registered. CONCLUSIONS: NS shows strong variable expressivity along the high genetic heterogeneity especially in distinct populations and ethnic groups. Also possibly unknown other causative genes may be exist. Obviously, more comprehensive and new technologies like NGS methods are the best choice for detection of molecular defects in patients for genotype, phenotype correlation and disease management.