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Retroviral vector-mediated expression of HoxB4 in hematopoietic cells using a novel coexpression strategy

Gene Therapy volume 8, pages 811–817 (2001)Cite this article

Abstract

Retroviral vector-mediated expression of the homeoboxgene, HoxB4, in hematopoietic cells has been reported to mediate a benign expansion of gene-modified hematopoietic stem and precursor cells in vivo. In the present study, we used a novel coexpression strategy for coordinated expression of HoxB4 along with a cytoplasmic protein from a retroviral vector. The novel coexpression strategy, based on cotranslational protein separation mediated by the 2A sequence of foot-and-mouth disease virus (FMDV), allows an indirect quantification of HoxB4 expression levels when inserting a reporter such as the enhanced green fluorescent protein (GFP) in the retroviral vector. Presence of the 2A sequence did not interfere with the correct subcellular localization of HoxB4 (nuclear) and GFP (cytoplasmic), nor with the titer of bicistronic vectors, and mediated functional long-term coexpression (at least 1 year) of GFP and HoxB4 after transplantation of transduced mouse bone marrow cells in mice.

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Acknowledgements

This work was supported by the ‘Bundesministerium für Bildung und Forschung’, BMBF 01KV9811 and BMBF 5008410. We thank Cordula Gruettner and Goekhan Arman-Kalcek for excellent technical assistance. We are also grateful to Keith Humphries and Michael Niepmann for providing us with the cDNAs of human HoxB4 and the FMDV-OK1 IRES, respectively.

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Author notes

  1. B Vogt

    Present address: Genethor GmbH, Robert-Roessle-Strasse 10, Berlin, D-13125, Germany

  2. M Ryan

    Present address: Division of Biomedical Sciences, University of St Andrews, School of Biology, Biomedical Sciences Building, North Haugh, St Andrews, UK

  3. C Baum

    Present address: Department Hematology and Oncology, Medizinische Hochschule, Carl-Neuberg-Strasse 1, Hannover, 30625, Germany

Authors and Affiliations

  1. Heinrich-Pette-Institut for Experimental Virology and Immunology, Martinistrasse 52, Hamburg, D-20251, Germany

    H Klump, B Schiedlmeier, B Vogt, M Ryan, W Ostertag & C Baum

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Klump, H., Schiedlmeier, B., Vogt, B. et al. Retroviral vector-mediated expression of HoxB4 in hematopoietic cells using a novel coexpression strategy. Gene Ther 8, 811–817 (2001). https://doi.org/10.1038/sj.gt.3301447

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