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Transient Tcf3 Gene Repression by TALE-Transcription Factor Targeting

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Abstract

Transplantation of hematopoietic stem and progenitor cells (HSCs) i.e., self-renewing cells that retain multipotentiality, is now a widely performed therapy for many hematopoietic diseases. However, these cells are present in low number and are subject to replicative senescence after extraction; thus, the acquisition of sufficient numbers of cells for transplantation requires donors able to provide repetitive blood samples and/or methods of expanding cell numbers without disturbing cell multipotentiality. Previous studies have shown that HSCs maintain their multipotentiality and self-renewal activity if TCF3 transcription function is blocked under B cell differentiating conditions. Taking advantage of this finding to devise a new approach to HSC expansion in vitro, we constructed an episomal expression vector that specifically targets and transiently represses the TCF3 gene. This consisted of a vector encoding a transcription activator-like effector (TALE) fused to a Krüppel-associated box (KRAB) repressor. We showed that this TALE-KRAB vector repressed expression of an exogenous reporter gene in HEK293 and COS-7 cell lines and, more importantly, efficiently repressed endogenous TCF3 in a human B lymphoma cell line. These findings suggest that this vector can be used to maintain multipotentiality in HSC being subjected to a long-term expansion regimen prior to transplantation.

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Acknowledgments

This work was supported in part by Japan Society for the Promotion of Science (JSPS) KAKENHI, grant numbers 25860797, and Okayama Foundation for Science and Technology.

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Authors and Affiliations

  1. Division of Medical Bioengineering, Graduate School of Natural Science and Technology, Okayama University, Okayama, 700-8530, Japan

    Junko Masuda, Akifumi Mizutani, Hiroshi Murakami, Narumi Maeno, Tsukasa Shigehiro, Ayano Satoh, Akimasa Seno, Vaidyanath Arun, Tomonari Kasai & Masaharu Seno

  2. Mucosal Immunity Section, Laboratory of Host Defenses, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, 20892, USA

    Junko Masuda, Warren Strober, Atsushi Kitani & Ivan J. Fuss

  3. Laboratory for Lymphocyte Development, RIKEN Research Center for Allergy and Immunology, Yokohama, 230-0045, Japan

    Hiroshi Kawamoto, Tomokatsu Ikawa & Yoshimoto Katsura

  4. Department of Immunology, Institute for Frontier Medical Sciences, Kyoto University, Kyoto, 606-8507, Japan

    Hiroshi Kawamoto

  5. Department of Oral Biochemistry, Asahi University School of Dentistry, Hozumi 1851, Gifu, 501-0296, Japan

    Eiji Takayama

  6. Laboratory for Immune Regeneration, RIKEN Center for Integrative Medical Sciences, Yokohama, 230-0045, Japan

    Tomokatsu Ikawa

  7. Division of Cell Regeneration and Transplantation, Advanced Medical Research Center, School of Medicine, Nihon University, Tokyo, 173-8610, Japan

    Yoshimoto Katsura

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  1. Junko Masuda
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  2. Hiroshi Kawamoto
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Correspondence to Junko Masuda.

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Masuda, J., Kawamoto, H., Strober, W. et al. Transient Tcf3 Gene Repression by TALE-Transcription Factor Targeting. Appl Biochem Biotechnol 180, 1559–1573 (2016). https://doi.org/10.1007/s12010-016-2187-4

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