sodium-dodecyl-sulfate and Elliptocytosis--Hereditary

Using clinical, morphological, genetic, and biochemical criteria, we studied ten white and North African families with hereditary elliptocytosis (HE). In four families, elliptocytic individuals displayed a highly significant reduction of band 4.1, which was recorded using two electrophoretic procedures. The 4.1a/4.1b ratio was also significantly reduced, as is usually observed in suspensions enriched in young red cells. This form of HE was invariably associated with the following characteristics: absence of clinical signs; numerous, smooth and well-elongated elliptocytes; dominant transmission; and, when investigated, normal osmotic fragility. Its frequency, among all forms of HE, is about one third as a first estimate, at least in whites and North Africans. In the other six families studied, elliptocytic subjects presented normal 4.1 bands. Again, the 4.1a/4.1b ratio was decreased, reflecting the red cell age-dependent changes in these two components. In three of these families, elliptocytosis was accompanied by clinical signs of variable intensity, and the mode of inheritance could not be unequivocally determined. Therefore, HE with a partially reduced band 4.1 defines a homogeneous variety of HE that can be isolated from other forms of HE. We suggest that it be termed the 4.1 (-) trait, so as to correspond with a previously proposed terminology.

Topics: Adolescent; Adult; Blood Proteins; Child; Child, Preschool; Cytoskeletal Proteins; Electrophoresis, Polyacrylamide Gel; Elliptocytosis, Hereditary; Erythrocyte Membrane; Female; Genetic Carrier Screening; Humans; Infant; Male; Membrane Proteins; Middle Aged; Neuropeptides; Pedigree; Sodium Dodecyl Sulfate

The sialoglycoproteins (glycophorins) in human red cell membranes of rare individuals lacking totally (Ge-1,-2,-3 phenotype) or partially (Ge-1,-2,3 phenotype) the Gerbich (Ge) blood group antigens and two Ge-1,-2,-3 heterozygotes were studied by dodecylsulfate polyacrylamide gel electrophoretic techniques. Two sialoglycoproteins (components D and E) were not detectable in the membranes from the homozygotes and found to be decreased by about 50% in those from the heterozygotes. Ge--1,-2,-3 and Ge-1,-2,3 cells were found to contain a 'new' component (mol. masses about 29 and 30 kDa, respectively) possibly representing a D/E hybrid molecule. This sialoglycoprotein was not detectable in membranes from the Ge-1,-2,-3 heterozygotes, suggesting that the Ge-1,-2,-3 phenotype may be caused by at least two different alleles at the Ge blood group antigen locus. Hemagglutination or hemagglutination inhibition tests involving anti-Ge 1,2,3 and -Ge 1,2 as well as native and enzyme-treated normal red cells (phenotype Ge 1,2,3) or membrane and sialoglycoprotein fractions from normal erythrocytes indicate that the receptors of these sera are located within the glycosylated domain(s) of the D and/or E sialoglycoprotein(s). Our data suggest that the Ge locus encodes the polypeptide sequences of the D and E sialoglycoproteins.

Topics: Blood Group Antigens; Densitometry; Electrophoresis; Elliptocytosis, Hereditary; Erythrocyte Membrane; Hemagglutination Inhibition Tests; Humans; Isoantigens; Rosaniline Dyes; Sialoglycoproteins; Sodium Dodecyl Sulfate

Two molecular defects involving the spectrin heterodimer (SpD) contact site of the alpha chain (the alpha I domain) were previously identified using limited tryptic digestion followed by two-dimensional isoelectric focusing/sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Both are characterized by atypical peptide maps which reveal a marked decrease of the 80,000-dalton alpha I domain and a formation of new major peptides of either 74,000 (Sp alpha I/74) or 46,000 (Sp alpha I/46) daltons. We now report a third variant of the spectrin alpha chain, designated Sp alpha I/65, in three unrelated black families. In all three probands, the percentage of SpD in the low ionic strength (O degrees C) membrane extracts was increased to 19% to 32%. One- and two-dimensional electrophoretic separations of limited tryptic digests of spectrin from all three probands revealed a decrease of the alpha I domain of spectrin and the concomitant appearance of peptides at 65,000 daltons and isoelectric points ranging from 5.2 to 5.3. The abnormal 65,000-dalton peptides could be stained with an antiserum which had been raised against the alpha I domain, indicating that it was derived from the alpha I domain.

Topics: Adult; Electrophoresis, Polyacrylamide Gel; Elliptocytosis, Hereditary; Female; Genetic Variation; Humans; Infant; Isoelectric Focusing; Macromolecular Substances; Middle Aged; Peptides; Sodium Dodecyl Sulfate; Spectrin

A family comprising three patients (a mother and two children) with mild hereditary elliptocytosis was studied. Each patient had prominent elliptocytosis, reduced red cell deformability, and normal erythrocyte thermal sensitivity. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of the erythrocyte membranes in each patient showed decreased levels of band 4.1 (approximately half of the normal value) and the presence of an additional band migrating below protein band 4.2. This additional band was shown to derive from protein 4.1. Comparative partial proteolytic mapping of protein 4.1 and the additional band revealed a number of common peptides. Enzyme-linked immunoelectrotransfer blots of the patients' erythrocyte membranes using a monoclonal antibody to protein 4.1 revealed that, in addition to protein 4.1, two other bands below protein 4.2 were stained; one of these bands migrated in the same position as the additional band detected in the Coomassie Blue-stained gels. Immunoblotting of the patients' whole cells using the antibody to protein 4.1 revealed that this altered band 4.1 occurred as such in the intact red cell. SDS-PAGE of protein 4.1 purified from one patient showed the presence of two lower molecular weight bands below protein 4.1; the lower band migrated in the same position as the additional band found on SDS-PAGE of the patients' erythrocyte membranes. The patient's purified protein 4.1 displayed a decrease of about 40% in the binding activity with crude spectrin extracted from normal controls. Spectrin-spectrin interactions were normal in the three patients. The additional band present in the patients' red cell membranes probably represents a proteolytic degradation product. This alteration, present both in whole cells and isolated membranes, might affect the intact cells in vivo. We suggest that the patients' erythrocyte membrane instability may be related to the presence of an abnormal protein 4.1 whose modulatory influence on the spectrin-actin interaction in the skeleton is defective.

Topics: Adult; Antibodies, Monoclonal; Blood Proteins; Cytoskeletal Proteins; Electrophoresis, Polyacrylamide Gel; Elliptocytosis, Hereditary; Erythrocyte Membrane; Female; Genetic Variation; Humans; Male; Membrane Proteins; Middle Aged; Neuropeptides; Sodium Dodecyl Sulfate; Spectrin