A simple, versatile and efficient method to genetically modify human monocyte-derived dendritic cells with HIV-1–derived lentiviral vectors
Lentiviral vectors derived from the human immunodeficiency type 1 virus (HIV-1 LV) are among the finest tools available today for the genetic modification of human monocyte-derived dendritic cells (MDDCs). However, this process is largely inefficient because MDDCs show a strong resistance to HIV-1 t...
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| Veröffentlicht in: | Nature protocols 2011-06, Vol.6 (6), p.806-816 |
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| creator | Berger, Grégory Durand, Stéphanie Goujon, Caroline Nguyen, Xuan-Nhi Cordeil, Stéphanie Darlix, Jean-Luc Cimarelli, Andrea |
| description | Lentiviral vectors derived from the human immunodeficiency type 1 virus (HIV-1 LV) are among the finest tools available today for the genetic modification of human monocyte-derived dendritic cells (MDDCs). However, this process is largely inefficient because MDDCs show a strong resistance to HIV-1 transduction. Here we describe a step-by-step protocol from the production of LVs to cell transduction that allows the efficient genetic modification of MDDCs. This protocol can be completed in 23 d from the initial phase of LV production to the final analysis of the results of MDDC transduction. The method relies on the simultaneous addition of HIV-1 LVs along with noninfectious virion-like particles carrying Vpx, a nonstructural protein encoded by the simian immunodeficiency virus (Vpx-VLPs). When thus provided in target cells, Vpx exerts a strong positive effect on incoming LVs by counteracting the restriction present in MDDCs; accordingly, 100% of cells can be transduced with low viral inputs. Vpx-VLPs will improve the efficiency of LV-mediated transduction of MDDCs with vectors for both ectopic gene expression and depletion studies. |
| doi_str_mv | 10.1038/nprot.2011.327 |
| format | Article |
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However, this process is largely inefficient because MDDCs show a strong resistance to HIV-1 transduction. Here we describe a step-by-step protocol from the production of LVs to cell transduction that allows the efficient genetic modification of MDDCs. This protocol can be completed in 23 d from the initial phase of LV production to the final analysis of the results of MDDC transduction. The method relies on the simultaneous addition of HIV-1 LVs along with noninfectious virion-like particles carrying Vpx, a nonstructural protein encoded by the simian immunodeficiency virus (Vpx-VLPs). When thus provided in target cells, Vpx exerts a strong positive effect on incoming LVs by counteracting the restriction present in MDDCs; accordingly, 100% of cells can be transduced with low viral inputs. Vpx-VLPs will improve the efficiency of LV-mediated transduction of MDDCs with vectors for both ectopic gene expression and depletion studies.</description><identifier>ISSN: 1754-2189</identifier><identifier>ISSN: 1750-2799</identifier><identifier>EISSN: 1750-2799</identifier><identifier>DOI: 10.1038/nprot.2011.327</identifier><identifier>PMID: 21637200</identifier><language>eng</language><publisher>London: Nature Publishing Group UK</publisher><subject>631/1647/2300/1851 ; 631/250/2504/133 ; 631/326/596/2561 ; Analytical Chemistry ; Antigens ; Biological Techniques ; Biomedical and Life Sciences ; Cell Culture Techniques ; Cell Differentiation ; Computational Biology/Bioinformatics ; Dendritic cells ; Dendritic Cells - cytology ; Dendritic Cells - virology ; Gene expression ; Genetic aspects ; Genetic Engineering - methods ; Genetic Vectors ; Genetically modified organisms ; HIV-1 - genetics ; Human immunodeficiency virus 1 ; Humans ; Immune system ; Infections ; Lentivirus ; Lentivirus - genetics ; Life Sciences ; Microarrays ; Microbiology and Parasitology ; Monocytes - cytology ; Organic Chemistry ; Physiological aspects ; Polypeptides ; Proteins ; Protocol ; Simian immunodeficiency virus ; Transduction, Genetic - methods ; Vectors (Biology) ; Viral Regulatory and Accessory Proteins - genetics ; Virion - genetics ; Virology ; Viruses</subject><ispartof>Nature protocols, 2011-06, Vol.6 (6), p.806-816</ispartof><rights>Springer Nature Limited 2011</rights><rights>COPYRIGHT 2011 Nature Publishing Group</rights><rights>Copyright Nature Publishing Group Jun 2011</rights><rights>Distributed under a Creative Commons Attribution 4.0 International License</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c528t-5d499196229d64a351234c2221e58b0de9984ca86d90f11a9937d97928a8e05c3</citedby><cites>FETCH-LOGICAL-c528t-5d499196229d64a351234c2221e58b0de9984ca86d90f11a9937d97928a8e05c3</cites><orcidid>0000-0001-8571-1108 ; 0000-0001-6461-7294 ; 0000-0003-0892-0488</orcidid></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://link.springer.com/content/pdf/10.1038/nprot.2011.327$$EPDF$$P50$$Gspringer$$H</linktopdf><linktohtml>$$Uhttps://link.springer.com/10.1038/nprot.2011.327$$EHTML$$P50$$Gspringer$$H</linktohtml><link.rule.ids>230,314,780,784,885,27924,27925,41488,42557,51319</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/21637200$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink><backlink>$$Uhttps://hal.science/hal-02322823$$DView record in HAL$$Hfree_for_read</backlink></links><search><creatorcontrib>Berger, Grégory</creatorcontrib><creatorcontrib>Durand, Stéphanie</creatorcontrib><creatorcontrib>Goujon, Caroline</creatorcontrib><creatorcontrib>Nguyen, Xuan-Nhi</creatorcontrib><creatorcontrib>Cordeil, Stéphanie</creatorcontrib><creatorcontrib>Darlix, Jean-Luc</creatorcontrib><creatorcontrib>Cimarelli, Andrea</creatorcontrib><title>A simple, versatile and efficient method to genetically modify human monocyte-derived dendritic cells with HIV-1–derived lentiviral vectors</title><title>Nature protocols</title><addtitle>Nat Protoc</addtitle><addtitle>Nat Protoc</addtitle><description>Lentiviral vectors derived from the human immunodeficiency type 1 virus (HIV-1 LV) are among the finest tools available today for the genetic modification of human monocyte-derived dendritic cells (MDDCs). 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Academic</collection><collection>Virology and AIDS Abstracts</collection><collection>Hyper Article en Ligne (HAL)</collection><jtitle>Nature protocols</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Berger, Grégory</au><au>Durand, Stéphanie</au><au>Goujon, Caroline</au><au>Nguyen, Xuan-Nhi</au><au>Cordeil, Stéphanie</au><au>Darlix, Jean-Luc</au><au>Cimarelli, Andrea</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>A simple, versatile and efficient method to genetically modify human monocyte-derived dendritic cells with HIV-1–derived lentiviral vectors</atitle><jtitle>Nature protocols</jtitle><stitle>Nat Protoc</stitle><addtitle>Nat Protoc</addtitle><date>2011-06</date><risdate>2011</risdate><volume>6</volume><issue>6</issue><spage>806</spage><epage>816</epage><pages>806-816</pages><issn>1754-2189</issn><issn>1750-2799</issn><eissn>1750-2799</eissn><abstract>Lentiviral vectors derived from the human immunodeficiency type 1 virus (HIV-1 LV) are among the finest tools available today for the genetic modification of human monocyte-derived dendritic cells (MDDCs). However, this process is largely inefficient because MDDCs show a strong resistance to HIV-1 transduction. Here we describe a step-by-step protocol from the production of LVs to cell transduction that allows the efficient genetic modification of MDDCs. This protocol can be completed in 23 d from the initial phase of LV production to the final analysis of the results of MDDC transduction. The method relies on the simultaneous addition of HIV-1 LVs along with noninfectious virion-like particles carrying Vpx, a nonstructural protein encoded by the simian immunodeficiency virus (Vpx-VLPs). When thus provided in target cells, Vpx exerts a strong positive effect on incoming LVs by counteracting the restriction present in MDDCs; accordingly, 100% of cells can be transduced with low viral inputs. Vpx-VLPs will improve the efficiency of LV-mediated transduction of MDDCs with vectors for both ectopic gene expression and depletion studies.</abstract><cop>London</cop><pub>Nature Publishing Group UK</pub><pmid>21637200</pmid><doi>10.1038/nprot.2011.327</doi><tpages>11</tpages><orcidid>https://orcid.org/0000-0001-8571-1108</orcidid><orcidid>https://orcid.org/0000-0001-6461-7294</orcidid><orcidid>https://orcid.org/0000-0003-0892-0488</orcidid></addata></record> |
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| subjects | 631/1647/2300/1851 631/250/2504/133 631/326/596/2561 Analytical Chemistry Antigens Biological Techniques Biomedical and Life Sciences Cell Culture Techniques Cell Differentiation Computational Biology/Bioinformatics Dendritic cells Dendritic Cells - cytology Dendritic Cells - virology Gene expression Genetic aspects Genetic Engineering - methods Genetic Vectors Genetically modified organisms HIV-1 - genetics Human immunodeficiency virus 1 Humans Immune system Infections Lentivirus Lentivirus - genetics Life Sciences Microarrays Microbiology and Parasitology Monocytes - cytology Organic Chemistry Physiological aspects Polypeptides Proteins Protocol Simian immunodeficiency virus Transduction, Genetic - methods Vectors (Biology) Viral Regulatory and Accessory Proteins - genetics Virion - genetics Virology Viruses |
| title | A simple, versatile and efficient method to genetically modify human monocyte-derived dendritic cells with HIV-1–derived lentiviral vectors |
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