Molecular Heterogeneity of Hereditary Pyropoikilocytosis: Identification of a Second Variant of the Spectrin α-Subunit

In hereditary pyropoikilocytosis (HPP), the red cell membrane skeletons exhibit a mechanical instability that can be correlated to defective self-association of spectrin heterodimers. To determine the underlying molecular defect, we have subjected HPP spectrin to limited tryptic digestion, followed by one- and two-dimensional separations of the peptides. Two of the HPP kindreds exhibited a marked decrease in 80,000-dalton peptide (previously identified as the spectrin dimer–dimer contact domain of the a-subunit) and a concomitant increase of the 74,000-dalton polypeptide (presumably derived from the 80,000-dalton domain) and a decrease in a 22,000-dalton polypeptide. We now report tryptic digests of two other HPP kindred that are characterized by a decrease or complete absence of the 80,000-dalton tryptic fragment, with a concomitant increase in fragments at 46,000 and 17,000 daltons. The 46,000-dalton fragment separated into multiple spots on isoelectric focusing, ranging in isoelectric point from 5.25 to 5.35, and the 17,000-dalton fragment focused to a single spot at 5.4. Minor fragments at 56,000 and 22,000 daltons were also decreased, while a 38,000-dalton fragment increased. Limited tryptic digestion of the separated α- and β-subunits revealed that the 74,000-dalton fragment in the first group of patients and the 46,000-dalton fragment in the second group of patients were derived from the α-subunit. Both subtypes exhibited a similar defect of spectrin self-association, with 30%-38% of spectrin dimers in 0°C extracts. The results indicate that at least two distinct forms of structurally defective spectrin may give rise to the clinical presentation of HPP.