A novel ENU-generated truncation mutation lacking the spectrin-binding and C-terminal regulatory domains of Ank1 models severe hemolytic hereditary spherocytosis

Exp Hematol. 2011 Mar;39(3):305-20, 320.e1-2. doi: 10.1016/j.exphem.2010.12.009. Epub 2010 Dec 28.


Objective: Hereditary spherocytosis (HS) is a heterogeneous group of spontaneously arising and inherited red blood cell disorders ranging from very mild subclinical cases to severe and life-threatening cases, with symptoms linked directly to the severity of the mutation at the molecular level. We investigated a novel mouse model in which the heterozygotes present with the diagnostic hallmarks of mild HS and surviving homozygotes phenocopy severe hemolytic HS.

Materials and methods: We used N-ethyl-N-nitrosourea mutagenesis to generate random point mutations in the mouse genome and a dominant screen to identify mouse models of human hematopoietic disease. Gene mapping of the HS strain revealed a unique in-frame nonsense mutation arising from a single base transversion in exon 27 of Ank1 (strain designation: Ank1(E924X)). Employing conventional hematopoietic, pathological, biochemical, and cell biology assays, we characterized heterozygous and homozygous Ank1(E924X) mice at the biochemical, cellular, and pathophysiological levels.

Results: Although Ank1(E924X/E924X) red blood cell ghosts lack abundant full-length ankyrin-1 isoforms, N-terminal epitope ankyrin-1 antibodies reveal a band consistent with the theoretical size of a truncated mutant ankyrin-1. Using domain-specific antibodies, we further show that this protein lacks both a spectrin-binding domain and a C-terminal regulatory domain. Finally, using antisera that detect C-terminal residues of the products of alternative Ank1 transcripts, we find unique immunoreactive bands not observed in red blood cell ghosts from wild-type or Ank1(E924X) heterozygous mice, including a band similar in size to full-length ankyrin-1.

Conclusions: The Ank1(E924X) strain provides a novel tool to study Ank1 and model HS.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alkylating Agents / adverse effects*
  • Alkylating Agents / pharmacology
  • Amino Acid Sequence
  • Animals
  • Ankyrins* / genetics
  • Ankyrins* / metabolism
  • Codon, Nonsense*
  • Disease Models, Animal
  • Erythrocytes / metabolism*
  • Ethylnitrosourea / adverse effects*
  • Ethylnitrosourea / pharmacology
  • Female
  • Humans
  • Male
  • Mice
  • Mice, Mutant Strains
  • Protein Binding
  • Sequence Deletion
  • Spherocytosis, Hereditary* / chemically induced
  • Spherocytosis, Hereditary* / genetics
  • Spherocytosis, Hereditary* / metabolism


  • Alkylating Agents
  • Ank1 protein, mouse
  • Ankyrins
  • Codon, Nonsense
  • Ethylnitrosourea