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Vpx-independent lentiviral transduction and shRNA-mediated protein knock-down in monocyte-derived dendritic cells

Wojciech Witkowski (UGent) , JOLIEN VERMEIRE (UGent) , Alessia Landi (UGent) , Evelien Naessens (UGent) , Hanne Vanderstraeten (UGent) , Hans Nauwynck (UGent) , Herman Favoreel (UGent) and Bruno Verhasselt (UGent)
(2015) PLOS ONE. 10(7).
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Abstract
The function of dendritic cells (DCs) in the immune system is based on their ability to sense and present foreign antigens. Powerful tools to research DC function and to apply in cell-based immunotherapy are either silencing or overexpression of genes achieved by lentiviral transduction. To date, efficient lentiviral transduction of DCs or their monocyte derived counterparts (MDDCs) required high multiplicity of infection (MOI) or the exposure to the HIV-2/SIV protein Vpx to degrade viral restriction factor SAM domain and HD domain-containing protein 1 (SAMHD1). Here we present a Vpx-independent method for efficient (>95%) transduction of MDDCs at lower MOI. The protocol can be used both for ectopic gene expression and knock-down. Introducing shRNA targeting viral entry receptor CD4 and restriction factor SAMHD1 into MDDCs resulted in down-regulation of targeted proteins and, consequently, expected impact on HIV infection. This protocol for MDDCs transduction is robust and free of the potential risk arising from the use of Vpx which creates a virus infection-prone environment, potentially dangerous in clinical setting.
Keywords
MACROPHAGES, PATHOGENESIS, SAMHD1 RESTRICTS, HIV-1 RESTRICTION, IMMUNODEFICIENCY-VIRUS, REPLICATION, ACTIVATION, INFECTION, IMMUNITY

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Chicago
Witkowski, Wojciech, JOLIEN VERMEIRE, Alessia Landi, Evelien Naessens, Hanne Vanderstraeten, Hans Nauwynck, Herman Favoreel, and Bruno Verhasselt. 2015. “Vpx-independent Lentiviral Transduction and shRNA-mediated Protein Knock-down in Monocyte-derived Dendritic Cells.” Plos One 10 (7).
APA
Witkowski, W., VERMEIRE, J., Landi, A., Naessens, E., Vanderstraeten, H., Nauwynck, H., Favoreel, H., et al. (2015). Vpx-independent lentiviral transduction and shRNA-mediated protein knock-down in monocyte-derived dendritic cells. PLOS ONE, 10(7).
Vancouver
1.
Witkowski W, VERMEIRE J, Landi A, Naessens E, Vanderstraeten H, Nauwynck H, et al. Vpx-independent lentiviral transduction and shRNA-mediated protein knock-down in monocyte-derived dendritic cells. PLOS ONE. 2015;10(7).
MLA
Witkowski, Wojciech, JOLIEN VERMEIRE, Alessia Landi, et al. “Vpx-independent Lentiviral Transduction and shRNA-mediated Protein Knock-down in Monocyte-derived Dendritic Cells.” PLOS ONE 10.7 (2015): n. pag. Print.
@article{6960781,
  abstract     = {The function of dendritic cells (DCs) in the immune system is based on their ability to sense and present foreign antigens. Powerful tools to research DC function and to apply in cell-based immunotherapy are either silencing or overexpression of genes achieved by lentiviral transduction. To date, efficient lentiviral transduction of DCs or their monocyte derived counterparts (MDDCs) required high multiplicity of infection (MOI) or the exposure to the HIV-2/SIV protein Vpx to degrade viral restriction factor SAM domain and HD domain-containing protein 1 (SAMHD1). Here we present a Vpx-independent method for efficient ({\textrangle}95\%) transduction of MDDCs at lower MOI. The protocol can be used both for ectopic gene expression and knock-down. Introducing shRNA targeting viral entry receptor CD4 and restriction factor SAMHD1 into MDDCs resulted in down-regulation of targeted proteins and, consequently, expected impact on HIV infection. This protocol for MDDCs transduction is robust and free of the potential risk arising from the use of Vpx which creates a virus infection-prone environment, potentially dangerous in clinical setting.},
  articleno    = {e0133651},
  author       = {Witkowski, Wojciech and VERMEIRE, JOLIEN and Landi, Alessia and Naessens, Evelien and Vanderstraeten, Hanne and Nauwynck, Hans and Favoreel, Herman and Verhasselt, Bruno},
  issn         = {1932-6203},
  journal      = {PLOS ONE},
  keyword      = {MACROPHAGES,PATHOGENESIS,SAMHD1 RESTRICTS,HIV-1 RESTRICTION,IMMUNODEFICIENCY-VIRUS,REPLICATION,ACTIVATION,INFECTION,IMMUNITY},
  language     = {eng},
  number       = {7},
  pages        = {12},
  title        = {Vpx-independent lentiviral transduction and shRNA-mediated protein knock-down in monocyte-derived dendritic cells},
  url          = {http://dx.doi.org/10.1371/journal.pone.0133651},
  volume       = {10},
  year         = {2015},
}

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