An insertional frameshift mutation of the β-spectrin gene associated with elliptocytosis in spectrin nice (β220/216)

Abstract

Spectrin Nice (β220/216) is a spectrin variant associated with a shortened β chain found in a patient with elliptocytosis. The shortened β chain (β′ chain) appeared as an additional band of approximately 216 Kd on sodium dodecyl sulfate-polyacrylamide gel electrophoresis and was defective in its ability to be phosphorylated. There were increased amounts of spectrin dimers in crude spectrin extracts from the propositus and the association constant of spectrin dimer self-association was decreased. There was an associated increase of the ol 74-Kd fragment from the α chain after partial trypic digestion of spectrin. To identify the underlying molecular defect, we analyzed cDNA for β spectrin obtained by polymerase chain reaction amplification of reverse-transcribed reticulocyte messenger RNA from peripheral blood of the propositus. DNA sequencing of individual as well as pooled subclones showed that two extra bases (GA) are inserted in codon no. 2046 in one allele of the β-spectrin gene. The insertion results in a frameshift mutation and generates an aberrant C-terminus truncated by about 4 Kd, consistent with the estimated size of the β′ chain observed. By allele-specific oligonucleotide hybridization, the insertion was shown to be present in the propositus and absent in his parents, confirming a previous proposal that it is a de novo mutation. The determination of the location of the mutation in spectrin Nice points to specific regions of the β-spectrin chain where phosphorylation may occur. A model is proposed to describe the interaction between the α- and β-spectrin chains and to explain the effects of the mutation found in spectrin Nice on the trypsin digestion pattern of its associated α chain. © 1991 by The American Society of Hematology.