Two Novel Mutations (HBG1: c.-250C>T and HBG2: c.-250C>T) Associated With Hereditary Persistence of Fetal Hemoglobin

Mutations within the promoters of either of the γ-globin genes [^Gγ (HBG1) and ^Aγ (HBG2)] lead to variably increased levels of fetal hemoglobin (Hb) (Hb F, α2γ2) in the syndrome of hereditary persistence of fetal Hb (HPFH). Carriers of such mutations are clinically asymptomatic and the mutations are usually detected as part of routine screening or family studies. We describe two new nondeletional HPFH mutations, both C>T substitutions at position c.-250, one in the HBG1 and the other in the HBG2 globin gene promoters.     

Intermediate hyperhomocysteinaemia and compound heterozygosity for the common variant c.677C>T and a MTHFR gene mutation

Summary Methylenetetrahydrofolate reductase (MTHFR) is a key enzyme in the regulation of plasma homocysteine levels. MTHFR deficiency, an autosomal recessive disorder, results in homocystinuria and hypomethioninaemia and presents with highly variable symptoms affecting many organs but predominantly the central nervous system. The common polymorphism of the MTHFR gene, c.677C>T, a known risk factor for elevated plasma homocysteine levels, occurs frequently in the caucasian population. In this study we investigated three subjects with moderate hyperhomocysteinaemia (total plasma homocysteine 72 μmol/L in case 1 and 90 μmol/L in case 3, total non-protein-bound homocysteine 144–186 μmol/L in case 2) but different clinical presentation with no symptoms in case 1, muscle weakness at 17 years of age in case 2, and syncopes and cerebral convulsions at 18 years of age in case 3. Each subject was compound heterozygous for the c.677C>T polymorphism and a novel mutation of the MTHFR gene (case 1: c.883G>A [p.D291N]; case 2: c.1552_c.1553delGA [p.E514fsX536]; case 3: c.616C>T [p.P202S]). Moderately decreased fibroblast MTHFR activity was associated with severely reduced affinity for NADPH and increased sensitivity to inhibition by S-adenosylmethionine (AdoMet) in case 2, and with mild FAD responsiveness in case 3. In case 1, fibroblast MTHFR activity was normal but the sensitivity to inhibition by AdoMet was slightly reduced. This study indicates that the sequence alteration c.677C>T combined with severe MTHFR mutations in compound heterozygous state may lead to moderate biochemical and clinical abnormalities exceeding those attributed to the c.677TT genotype and might require in addition to folate substitution further therapy to normalize homocysteine levels. Electronic Supplementary Material The online version of this article (doi:) contains supplementary material, which is available to authorized users. Online citation: JIMD Short Report #048 (2007) Online     

A novel DO null allele with a c.268C>T (p.Gln90Stop) mutation in Japanese

The Dombrock blood group system consists of two antithetical antigens, Do^a (DO1) and Do^b (DO2), and seven high‐prevalence antigens, Gy^a (DO3), Hy (DO4), Jo^a (DO5), DOYA (DO6), DOMR (DO7), DOLG (DO8) and DOLC (DO9). Do^a/Do^b polymorphism is associated with c.793A>G (p.Asn265Asp) in exon 2 of the DO (ART4) gene, and the corresponding alleles are named DO*01 and DO*02. The rare Do_null or Gy(a?) phenotype lacks all Dombrock antigens, and the DO null alleles vary with both DO*01 and DO*02 backgrounds. We report a novel DO null allele, which has a c.268C>T (p.Gln90Stop) nonsense mutation with a DO*02 background identified from four unrelated Gy(a?) Japanese individuals.     

Association of insulin sensitivity and glucose tolerance with the c.825C>T variant of the G protein beta-3 subunit gene

The risk of macrovascular complications of diabetes mellitus is greatly enhanced by the presence of high blood pressure. In addition, hypertension and diabetes share insulin resistance as a common pathophysiological mechanism. Despite evidence for a common molecular genetic background of insulin resistance, glucose intolerance, and hypertension, few candidate genes have been shown to influence all of these features simultaneously. We examined the association of insulin sensitivity with the c.825C>T variant of the g-protein beta-3 subunit (GNB3), a candidate gene of hypertension, in families of Mexican-American hypertensive patients.

Methods

One hundred eighty subjects enrolled in a family study of Mexican-American hypertensive patients were recruited from hypertension clinics in Los Angeles. Subjects underwent pretreatment blood pressure recording, an oral glucose tolerance test, euglycemic hyperinsulinemic clamp, and anthropometric measurements. DNA from peripheral blood leukocytes was genotyped by polymerase chain reaction and restriction enzyme digest with BseD1 (GNB). Statistical analysis was performed by transmission disequilibrium testing.

Results

In carriers of the T-allele, blood glucose was significantly lower [(mean+S.D.) fasting: 96.7+22.9 vs. 106.7+51.7mg/dl, P=.009; oral glucose tolerance test (oGTT) 120 min: 131.7+48.7 vs. 137.8+64.9 mg/dl, P=.036], and insulin sensitivity was significantly higher (229.0+108.7 vs. 188.5+94.2 mg/kg per minute, P=.037) than in homozygous carriers of the C-allele. Blood pressure did not differ significantly between the phenotypes.

Conclusion

In a Mexican-American hypertensive population, we found evidence for higher insulin sensitivity in carriers of the T allele of the c.825C>T variant of GNB3.     

Hb Presbyterian (HBB: c.327C>G) in a Nicaraguan Family

Hemoglobin (Hb) is the protein responsible for oxygen transportation. It is a tetrameric protein comprising two α- and two β-globin subunits. In the literature, a large number of mutations in the α- and β-globin genes have been documented. Among these mutations, Hb Presbyterian (HBB: c.327?C>G), is a naturally occurring mutant exerting low oxygen affinity. The C to G exchange (AAC>AAG) at codon 108 of the β-globin gene results in the substitution of asparagine by lysine. Here, we document the identification of HBB: c.327?C>G in a 6-year-old female patient and her father from Nicaragua and Cuba, respectively. The presence of the abnormal Hb was confirmed by cellulose acetate electrophoresis, high performance liquid chromatography (HPLC) and genomic DNA sequencing. The β-globin gene sequences for both, father and daughter, disclosed the heterozygous mutation at codon 108 to be Hb Presbyterian or HBB: c.327?C>G. The mutant Hb was previously reported in four families from North America, Germany, Japan and Spain, respectively. This is the fifth family carrying HBB: c.327?C>G described to date and the first report from Latin America.     

“ A.T. 10 “ AND DIHYDROTACHYSTEROL

The effect of pure dihydrotachysterol (D.H.T.) was compared with that of the old “ A.T. 10 ” in eight patients with hypoparathyroidism, and it was found that the “ A.T. 10 ” available in 1963–1964 was equivalent in therapeutic activity to 0.1 mg. of D.H.T. per capsule (or 0.2 mg. of D.H.T. per millilitre). It is concluded that the new “ A.T. 10 ” (containing 0.25 mg. of D.H.T. per millilitre) is 25% stronger than the old, and that patients whose therapy is changed to the new preparation in the same dose (by volume) may occasionally develop hypercalcaemia as a result. The history of the various changes in the methods of assay of “ A.T. 10 ” is reviewed and the reasons why they were all fallacious are explained. As judged by reported maintenance doses, the “ A.T. 10 ” available in 1963–1964 was only one-third as potent in the treatment of hypoparathyroidism as that available in 1934–1942.     

A New Hemoglobin Variant: Hb Izmir [β86(F2)Ala→Val,GCC>GTC; HBB:c.260C>T]

We report a new hemoglobin (Hb) variant [β86(F2)Ala→Val; HBB:c.260C>T] that we have named Hb Izmir. We have identified Hb Izmir in a Turkish woman by ion exchange high performance liquid chromatography (HPLC) during a premarital screening program in the Aegean region of Turkey. The mother and sister of the proband also carried the same variant. Using direct sequencing, we have characterized this variant as resulting from a GCC>GTC replacement at codon 86 of the β-globin chain, corresponding to an Ala→Val amino acid substitution. In the heterozygote, the level of Hb Izmir ranged from 41.38 to 45.6%. All heterozygotes had a Hb A₂ level of less than 3.5%. Total blood count values were normal and there were no other clinical findings. Although its clinical significance is thus far unclear, Hb Izmir may be important in hemoglobinopathy screening programs.     

An apparently silent nucleotide substitution (c.7056C>T) in the von Willebrand factor gene is responsible for type 1 von Willebrand disease

Background

Nucleotide variations not changing protein sequences are considered silent mutations; accumulating data suggest that they can, however, be important in human diseases.

Design and Methods

We report an altered splicing process induced by a silent substitution (c.7056C>T) in the von Willebrand factor gene in a case of type 1 von Willebrand disease originally classified as lacking von Willebrand factor mutations.

Results

The c.7056C>T synonymous substitution introduces a new donor splice site within exon 41, leading to messenger RNA lacking nucleotides 7055-7081 (c.7055_7081del). The encoded von Willebrand factor protein is predicted to lack amino acids 2352-2360 in the B2 domain. The patient’s von Willebrand disease phenotype was characterized by reduced plasma and platelet von Willebrand factor, which was normal in function and multimer structure. In vitro expression studies demonstrated that co-transfection of equimolar c.7055_7081del and wild-type von Willebrand factor (mimicking the patient’s heterozygous state) induced a 50% lower von Willebrand factor secretion than the wild type, while almost no von Willebrand factor secretion was seen with the mutated von Willebrand factor alone. The secreted von Willebrand factor was structurally and functionally normal, suggesting that the c.7056C>T substitution behaves like a loss-of-function allele.

Conclusions

This is the first report of a synonymous von Willebrand factor substitution being responsible for von Willebrand disease. Our findings suggest the need to reconsider the role of von Willebrand factor polymorphisms in von Willebrand disease.