Transient Tcf3 Gene Repression by TALE-Transcription Factor Targeting

• Published in: Applied biochemistry and biotechnology Vol. 180; no. 8; pp. 1559 - 1573
• Main Authors: Masuda, Junko, Kawamoto, Hiroshi, Strober, Warren, Takayama, Eiji, Mizutani, Akifumi, Murakami, Hiroshi, Ikawa, Tomokatsu, Kitani, Atsushi, Maeno, Narumi, Shigehiro, Tsukasa, Satoh, Ayano, Seno, Akimasa, Arun, Vaidyanath, Kasai, Tomonari, Fuss, Ivan J., Katsura, Yoshimoto, Seno, Masaharu
• Format: Journal Article
• Language: English
• Published: New York Springer US 01.12.2016; Springer Nature B.V
• Subjects: Animals; B-lymphocytes; Basic Helix-Loop-Helix Transcription Factors - genetics; Biochemistry; Biotechnology; Blood; blood sampling; cell senescence; Cellular biology; Chemistry; Chemistry and Materials Science; Chlorocebus aethiops; COS Cells; Gene Deletion; Gene expression; Gene Targeting; Genes, Reporter; Green Fluorescent Proteins - metabolism; HEK293 Cells; Humans; Luciferases - metabolism; Luminescent Proteins - metabolism; Lymphoma; Plasmids - metabolism; Red Fluorescent Protein; reporter genes; Repressor Proteins - metabolism; Stem cells; therapeutics; transcription (genetics); Transcription Activator-Like Effectors - metabolism; Transfection; Transplantation; Transplants & implants; TCF3 (E2A); TALE technology; Artificial transcription factor
• ISSN: 0273-2289; 1559-0291
• DOI: 10.1007/s12010-016-2187-4

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.