Erythrocyte ankyrin promoter mutations associated with recessive hereditary spherocytosis cause significant abnormalities in ankyrin expression

Ankyrin defects are the most common cause of hereditary spherocytosis (HS). In several kindreds with recessive, ankyrin-deficient HS, mutations have b een identified in the ankyrin promoter that have been proposed to decrease ankyrin synthesis. We analyzed the effects of two mutations, - 108T to C an d - 108T to C in cis with - 153G to A, on ankyrin expression. No difference between wild type and mutant promoters was demonstrated in transfection or gel shift assays in vitro. Transgenic mice with a wild type ankyrin promot er linked to a human (A)gamma -globin gene expressed gamma -globin in 100% of erythrocytes in a copy number-dependent, position-independent manner. Tr ansgenic mice with the mutant -108 promoter demonstrated variegated gamma - globin expression, but showed copy number-dependent and position-independen t expression similar to wild type. Severe effects in ankyrin expression wer e seen in mice with the linked -108/-153 mutations. Three transgenic lines had undetectable levels of (A)gamma -globin mRNA, indicating position-depen dent expression, and four lines expressed significantly lower levels of (A) gamma -globin mRNA than wild type. Two of four expressing lines showed vari egated gamma -globin expression, and there was no correlation between trans gene copy number and RNA level, indicating copy number-independent expressi on. These data are the first demonstration of functional defects caused by HS-related, ankyrin gene promoter mutations.