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

Michael R. Hughes, Nicole Anderson, Steven Maltby, Justin Wong, Zorana Berberovic, Connie S. Birkenmeier, D. James Haddon, Kamal Garcha, Ann Flenniken, Lucy R. Osborne, S. Lee Adamson, Janet Rossant, Luanne L. Peters, Mark D. Minden, Robert F. Paulson, Chen Wang, Dwayne L. Barber, Kelly M. McNagny, William L. Stanford

Research output: Contribution to journalArticle

21 Scopus citations

Abstract

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: Ank1E924X). Employing conventional hematopoietic, pathological, biochemical, and cell biology assays, we characterized heterozygous and homozygous Ank1E924X mice at the biochemical, cellular, and pathophysiological levels. Results: Although Ank1E924X/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 Ank1E924X heterozygous mice, including a band similar in size to full-length ankyrin-1. Conclusions: The Ank1E924X strain provides a novel tool to study Ank1 and model HS.

Original languageEnglish (US)
Pages (from-to)305-320.e2
JournalExperimental Hematology
Volume39
Issue number3
DOIs
StatePublished - Mar 2011

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Hematology
  • Genetics
  • Cell Biology
  • Cancer Research

Fingerprint Dive into the research topics of 'A novel ENU-generated truncation mutation lacking the spectrin-binding and C-terminal regulatory domains of Ank1 models severe hemolytic hereditary spherocytosis'. Together they form a unique fingerprint.

  • Cite this

    Hughes, M. R., Anderson, N., Maltby, S., Wong, J., Berberovic, Z., Birkenmeier, C. S., Haddon, D. J., Garcha, K., Flenniken, A., Osborne, L. R., Adamson, S. L., Rossant, J., Peters, L. L., Minden, M. D., Paulson, R. F., Wang, C., Barber, D. L., McNagny, K. M., & Stanford, W. L. (2011). A novel ENU-generated truncation mutation lacking the spectrin-binding and C-terminal regulatory domains of Ank1 models severe hemolytic hereditary spherocytosis. Experimental Hematology, 39(3), 305-320.e2. https://doi.org/10.1016/j.exphem.2010.12.009