title: Erythrocyte ankyrin promoter mutations associated with recessive hereditary spherocytosis cause significant abnormalities in ankyrin expression.
abstract: Ankyrin defects are the most common cause of hereditary spherocytosis (HS). In several kindreds with recessive, ankyrin-deficient HS, mutations have been identified in the ankyrin promoter that have been proposed to decrease ankyrin synthesis. We analyzed the effects of two mutations, -108T to C and -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 promoter linked to a human (A)gamma-globin gene expressed gamma-globin in 100% of erythrocytes in a copy number-dependent, position-independent manner. Transgenic mice with the mutant -108 promoter demonstrated variegated gamma-globin expression, but showed copy number-dependent and position-independent expression similar to wild type. Severe effects in ankyrin expression were seen in mice with the linked -108/-153 mutations. Three transgenic lines had undetectable levels of (A)gamma-globin mRNA, indicating position-dependent expression, and four lines expressed significantly lower levels of (A)gamma-globin mRNA than wild type. Two of four expressing lines showed variegated gamma-globin expression, and there was no correlation between transgene copy number and RNA level, indicating copy number-independent expression. These data are the first demonstration of functional defects caused by HS-related, ankyrin gene promoter mutations.
authors.affiliation: Department of Pediatrics, Yale University School of Medicine, New Haven, Connecticut 06520, USA. patrick.gallagher@yale.edu
authors.forename: P G
authors.initials: PG
authors.lastname: Gallagher
authors.forename: D E
authors.initials: DE
authors.lastname: Sabatino
authors.forename: D S
authors.initials: DS
authors.lastname: Basseres
authors.forename: D M
authors.initials: DM
authors.lastname: Nilson
authors.forename: C
authors.initials: C
authors.lastname: Wong
authors.forename: A P
authors.initials: AP
authors.lastname: Cline
authors.forename: L J
authors.initials: LJ
authors.lastname: Garrett
authors.forename: D M
authors.initials: DM
authors.lastname: Bodine
mesh_terms: D000818:Animals; D017487:Ankyrins / Q000235:genetics*; D004247:DNA / Q000378:metabolism; D018628:Gene Dosage; D005914:Globins / Q000032:analysis / Q000235:genetics; D006801:Humans; D020014:K562 Cells; D051379:Mice; D008822:Mice, Transgenic; D009154:Mutation; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000032:analysis; D013103:Spherocytosis, Hereditary / Q000235:genetics*; D014162:Transfection
chemical_list: D017487:Ankyrins; D012333:RNA, Messenger; D005914:Globins; D004247:DNA