Abstract
Transduction of lymphoid progenitors with retroviral or lentiviral vectors is a powerful experimental strategy to tease out the role of a gene or pathway in T cell development via gain-of-function or loss-of-function strategies. Here we discuss different approaches to use this powerful technology, and present some protocols that we use to transduce murine HSCs, thymocytes, and lymphoid cell lines with these viral vectors.
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References
Hawley RG, Fong AZ, Burns BF et al (1992) Transplantable myeloproliferative disease induced in mice by an interleukin 6 retrovirus. J Exp Med 176:1149–1163
Swift S, Lorens J, Achacoso P et al (2001) Rapid production of retroviruses for efficient gene delivery to mammalian cells using 293T cell-based systems. Curr Protoc Immunol Chapter 10:Unit 10.17C
Burns JC, Friedmann T, Driever W et al (1993) Vesicular stomatitis virus G glycoprotein pseudotyped retroviral vectors: concentration to very high titer and efficient gene transfer into mammalian and nonmammalian cells. Proc Natl Acad Sci U S A 90:8033–8037
Mostoslavsky G, Kotton DN, Fabian AJ et al (2005) Efficiency of transduction of highly purified murine hematopoietic stem cells by lentiviral and oncoretroviral vectors under conditions of minimal in vitro manipulation. Mol Ther 11:932–940
Schambach A, Bohne J, Chandra S et al (2006) Equal potency of gammaretroviral and lentiviral SIN vectors for expression of O6-methylguanine-DNA methyltransferase in hematopoietic cells. Mol Ther 13:391–400
Barrette S, Douglas JL, Seidel NE et al (2000) Lentivirus-based vectors transduce mouse hematopoietic stem cells with similar efficiency to moloney murine leukemia virus-based vectors. Blood 96:3385–3391
Laurenti E, Barde I, Verp S et al (2010) Inducible gene and shRNA expression in resident hematopoietic stem cells in vivo. Stem Cells 28:1390–1398
Tiscornia G, Tergaonkar V, Galimi F et al (2004) CRE recombinase-inducible RNA interference mediated by lentiviral vectors. Proc Natl Acad Sci U S A 101:7347–7351
Heinonen JE, Mohamed AJ, Nore BF et al (2005) Inducible H1 promoter-driven lentiviral siRNA expression by Stuffer reporter deletion. Oligonucleotides 15:139–144
Koo BK, Stange DE, Sato T et al (2012) Controlled gene expression in primary Lgr5 organoid cultures. Nat Methods 9:81–83
Kowolik CM, Hu J, Yee JK (2001) Locus control region of the human CD2 gene in a lentivirus vector confers position-independent transgene expression. J Virol 75:4641–4648
Marodon G, Mouly E, Blair EJ et al (2003) Specific transgene expression in human and mouse CD4+ cells using lentiviral vectors with regulatory sequences from the CD4 gene. Blood 101:3416–3423
Lois C, Hong EJ, Pease S et al (2002) Germline transmission and tissue-specific expression of transgenes delivered by lentiviral vectors. Science 295:868–872
Romero Z, Torres S, Cobo M et al (2011) A tissue-specific, activation-inducible, lentiviral vector regulated by human CD40L proximal promoter sequences. Gene Ther 18:364–371
Kraunus J, Schaumann DH, Meyer J et al (2004) Self-inactivating retroviral vectors with improved RNA processing. Gene Ther 11:1568–1578
Schambach A, Mueller D, Galla M et al (2006) Overcoming promoter competition in packaging cells improves production of self-inactivating retroviral vectors. Gene Ther 13:1524–1533
Schmitt TM, Zuniga-Pflucker JC (2002) Induction of T cell development from hematopoietic progenitor cells by delta-like-1 in vitro. Immunity 17:749–756
Lerner C, Harrison DE (1990) 5-Fluorouracil spares hemopoietic stem cells responsible for long-term repopulation. Exp Hematol 18:114–118
Harrison DE, Lerner CP (1991) Most primitive hematopoietic stem cells are stimulated to cycle rapidly after treatment with 5-fluorouracil. Blood 78:1237–1240
Naviaux RK, Costanzi E, Haas M et al (1996) The pCL vector system: rapid production of helper-free, high-titer, recombinant retroviruses. J Virol 70:5701–5705
Pear WS, Miller JP, Xu L et al (1998) Efficient and rapid induction of a chronic myelogenous leukemia-like myeloproliferative disease in mice receiving P210 bcr/abl-transduced bone marrow. Blood 92:3780–3792
Lee NS, Dohjima T, Bauer G et al (2002) Expression of small interfering RNAs targeted against HIV-1 rev transcripts in human cells. Nat Biotechnol 20:500–555
Hernandez-Hoyos G, Alberola-Ila J (2005) Analysis of T-cell development by using short interfering RNA to knock down protein expression. Methods Enzymol 392:199–217
Paithankar KR, Prasad KS (1991) Precipitation of DNA by polyethylene glycol and ethanol. Nucleic Acids Res 19:1346
Kumar M, Keller B, Makalou N et al (2001) Systematic determination of the packaging limit of lentiviral vectors. Hum Gene Ther 12:1893–1905
Moritz T, Patel VP, Williams DA (1994) Bone marrow extracellular matrix molecules improve gene transfer into human hematopoietic cells via retroviral vectors. J Clin Invest 93:1451–1457
Tonks A, Tonks AJ, Pearn L et al (2005) Optimized retroviral transduction protocol which preserves the primitive subpopulation of human hematopoietic cells. Biotechnol Prog 21:953–958
Bain G, Quong MW, Soloff RS et al (1999) Thymocyte maturation is regulated by the activity of the helix-loop-helix protein, E47. J Exp Med 190:1605–1616
Gimferrer I, Hu T, Simmons A et al (2011) Regulation of GATA-3 expression during CD4 lineage differentiation. J Immunol 186:3892–3898
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Simmons, A., Alberola-Ila, J. (2016). Retroviral Transduction of T Cells and T Cell Precursors. In: Bosselut, R., S. Vacchio, M. (eds) T-Cell Development. Methods in Molecular Biology, vol 1323. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2809-5_8
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DOI: https://doi.org/10.1007/978-1-4939-2809-5_8
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-2808-8
Online ISBN: 978-1-4939-2809-5
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