title: All-in-One inducible lentiviral vector systems based on drug controlled FLP recombinase.
abstract: Site specific recombinases are frequently used as gene switches in transgenic animals where recombination is induced by drug treatment or by tissue specific recombinase expression. Alternatively, lentiviral gene transfer can be utilized for the genetic modification of a wide variety of cell types, albeit systems for tight control of transcriptional activity are scarce. Here, we combined lentiviral gene transfer and the development of a tightly drug-controlled FLP recombinase for the construction of "All-in-One" inducible gene expression systems. Tight control of FLP activity was achieved through N-terminal fusion with a FKBP12-derived conditional destruction domain and a C-terminal estrogen receptor binding domain making recombination dependent on the presence of Shield-1 and 4-hydroxytamoxifen. Exploiting the capacity of FLP to mediate excision and inversion, "All-in-One" lentiviral gene switch vector systems were generated where drug-induced recombination resulted in abrogation of FLP expression and subsequent overexpression or knockdown of the prototypical tumor suppressor phosphatase and tensin homolog PTEN. "All-in-One" vectors proved their functionality in a variety of hematopoietic cell lines, and primary murine bone marrow cells. Our new vector system thus combines the ease of lentiviral gene transfer and the power of site specific recombinases for analysis of gene function.
authors: Institute of Experimental Hematology, Hannover Medical School, Germany.
authors: Tobias
authors: T
authors: Maetzig
authors: Institute of Experimental Hematology, Hannover Medical School, Germany.
authors: Johannes
authors: J
authors: Kuehle
authors: Institute of Experimental Hematology, Hannover Medical School, Germany.
authors: Adrian
authors: A
authors: Schwarzer
authors: Institute of Experimental Hematology, Hannover Medical School, Germany.
authors: Soeren
authors: S
authors: Turan
authors: Institute of Experimental Hematology, Hannover Medical School, Germany.
authors: Michael
authors: M
authors: Rothe
authors: Department of Hematology, Hemostasis, Oncology and Stem Cell Transplantation, Hannover Medical School, Germany.
authors: Anuhar
authors: A
authors: Chaturvedi
authors: Institute of Experimental Hematology, Hannover Medical School, Germany.
authors: Michael
authors: M
authors: Morgan
authors: Institute of Experimental Hematology, Hannover Medical School, Germany.
authors: Teng Cheong
authors: TC
authors: Ha
authors: Department of Hematology, Hemostasis, Oncology and Stem Cell Transplantation, Hannover Medical School, Germany.
authors: Michael
authors: M
authors: Heuser
authors: Research Unit Gene Vectors, Helmholtz Zentrum München, German Research Center for Environmental Health, Munich, Germany; German Centre for Infection Research (DZIF), Partner Site Munich, Germany.
authors: Wolfgang
authors: W
authors: Hammerschmidt
authors: Institute of Experimental Hematology, Hannover Medical School, Germany.
authors: Christopher
authors: C
authors: Baum
authors: Institute of Experimental Hematology, Hannover Medical School, Germany; Division of Hematology/Oncology, Boston Children's Hospital, Harvard Medical School, Boston, USA. Electronic address: schambach.axel@mh-hannover.de.
authors: Axel
authors: A
authors: Schambach
mesh_terms: D000818:Animals; D001854:Bone Marrow Cells / Q000187:drug effects / Q000378:metabolism; D002460:Cell Line; D003062:Codon / Q000235:genetics; D004254:DNA Nucleotidyltransferases / Q000378:metabolism*; D016503:Drug Delivery Systems; D005260:Female; D055785:Gene Knockdown Techniques; D005822:Genetic Vectors / Q000378:metabolism*; D006801:Humans; D016086:Lentivirus / Q000235:genetics*; D051379:Mice; D008810:Mice, Inbred C57BL; D009025:Morpholines / Q000494:pharmacology*; D051059:PTEN Phosphohydrolase / Q000378:metabolism; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D022061:Tacrolimus Binding Protein 1A / Q000378:metabolism; D013629:Tamoxifen / Q000031:analogs & derivatives* / Q000494:pharmacology
chemical_list: D003062:Codon; D009025:Morpholines; D011993:Recombinant Fusion Proteins; C540198:Shield-1 compound; D013629:Tamoxifen; C016601:afimoxifene; D004254:DNA Nucleotidyltransferases; C045119:FLP recombinase; D051059:PTEN Phosphohydrolase; D022061:Tacrolimus Binding Protein 1A
keywords: ERT2; FKBP12; FLP recombinase; Inducible; Knockdown; Lentivirus