New α- and SIN γ-retrovectors for safe transduction and specific transgene expression in pancreatic β cell lines
Abstract Background Viral vectors are invaluable tools to transfer genes and/or regulatory sequences into differentiated cells such as pancreatic cells. To date, several kinds of viral vectors have been used to transduce different pancreatic cell types, including insulin-producing β cells. However,...
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| Format: | Article |
| Language: | English |
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BMC
2019-06-01
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| Series: | BMC Biotechnology |
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| Online Access: | http://link.springer.com/article/10.1186/s12896-019-0531-9 |
| _version_ | 1818333739345772544 |
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| author | Olivier Albagli Alicia Maugein Lukas Huijbregts Delphine Bredel Géraldine Carlier Patrick Martin Raphaël Scharfmann |
| author_facet | Olivier Albagli Alicia Maugein Lukas Huijbregts Delphine Bredel Géraldine Carlier Patrick Martin Raphaël Scharfmann |
| author_sort | Olivier Albagli |
| collection | DOAJ |
| description | Abstract Background Viral vectors are invaluable tools to transfer genes and/or regulatory sequences into differentiated cells such as pancreatic cells. To date, several kinds of viral vectors have been used to transduce different pancreatic cell types, including insulin-producing β cells. However, few studies have used vectors derived from « simple » retroviruses, such as avian α- or mouse γ-retroviruses, despite their high experimental convenience. Moreover, such vectors were never designed to specifically target transgene expression into β cells. Results We here describe two novel α- or SIN (Self-Inactivating) γ-retrovectors containing the RIP (Rat Insulin Promoter) as internal promoter. These two retrovectors are easily produced in standard BSL2 conditions, rapidly concentrated if needed, and harbor a large multiple cloning site. For the SIN γ-retrovector, either the VSV-G (pantropic) or the retroviral ecotropic (rodent specific) envelope was used. For the α-retrovector, we used the A type envelope, as its receptor, termed TVA, is only naturally present in avian cells and can efficiently be provided to mammalian β cells through either exogenous expression upon cDNA transfer or gesicle-mediated delivery of the protein. As expected, the transgenes cloned into the two RIP-containing retrovectors displayed a strong preferential expression in β over non-β cells compared to transgenes cloned in their non-RIP (CMV- or LTR-) regulated counterparts. We further show that RIP activity of both retrovectors mirrored fluctuations affecting endogenous INSULIN gene expression in human β cells. Finally, both α- and SIN γ-retrovectors were extremely poorly mobilized by the BXV1 xenotropic retrovirus, a common invader of human cells grown in immunodeficient mice, and, most notably, of human β cell lines. Conclusion Our novel α- and SIN γ-retrovectors are safe and convenient tools to stably and specifically express transgene(s) in mammalian β cells. Moreover, they both reproduce some regulatory patterns affecting INSULIN gene expression. Thus, they provide a helpful tool to both study the genetic control of β cell function and monitor changes in their differentiation status. |
| first_indexed | 2024-12-13T13:56:25Z |
| format | Article |
| id | doaj.art-d3b03597381245a7bec16a0ee70a5512 |
| institution | Directory Open Access Journal |
| issn | 1472-6750 |
| language | English |
| last_indexed | 2024-12-13T13:56:25Z |
| publishDate | 2019-06-01 |
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| series | BMC Biotechnology |
| spelling | doaj.art-d3b03597381245a7bec16a0ee70a55122022-12-21T23:42:53ZengBMCBMC Biotechnology1472-67502019-06-0119111810.1186/s12896-019-0531-9New α- and SIN γ-retrovectors for safe transduction and specific transgene expression in pancreatic β cell linesOlivier Albagli0Alicia Maugein1Lukas Huijbregts2Delphine Bredel3Géraldine Carlier4Patrick Martin5Raphaël Scharfmann6INSERM U1016, CNRS UMR8104, Institut Cochin, Université Paris DescartesINSERM U1016, CNRS UMR8104, Institut Cochin, Université Paris DescartesINSERM U1016, CNRS UMR8104, Institut Cochin, Université Paris DescartesINSERM U1016, CNRS UMR8104, Institut Cochin, Université Paris DescartesINSERM U1016, CNRS UMR8104, Institut Cochin, Université Paris DescartesUniversité Côte d’Azur, CNRS UMR7277 INSERM U1099, iBV (Institut de Biologie Valrose), Université Nice Sophia Antipolis, Bâtiment Sciences Naturelles; UFR SciencesINSERM U1016, CNRS UMR8104, Institut Cochin, Université Paris DescartesAbstract Background Viral vectors are invaluable tools to transfer genes and/or regulatory sequences into differentiated cells such as pancreatic cells. To date, several kinds of viral vectors have been used to transduce different pancreatic cell types, including insulin-producing β cells. However, few studies have used vectors derived from « simple » retroviruses, such as avian α- or mouse γ-retroviruses, despite their high experimental convenience. Moreover, such vectors were never designed to specifically target transgene expression into β cells. Results We here describe two novel α- or SIN (Self-Inactivating) γ-retrovectors containing the RIP (Rat Insulin Promoter) as internal promoter. These two retrovectors are easily produced in standard BSL2 conditions, rapidly concentrated if needed, and harbor a large multiple cloning site. For the SIN γ-retrovector, either the VSV-G (pantropic) or the retroviral ecotropic (rodent specific) envelope was used. For the α-retrovector, we used the A type envelope, as its receptor, termed TVA, is only naturally present in avian cells and can efficiently be provided to mammalian β cells through either exogenous expression upon cDNA transfer or gesicle-mediated delivery of the protein. As expected, the transgenes cloned into the two RIP-containing retrovectors displayed a strong preferential expression in β over non-β cells compared to transgenes cloned in their non-RIP (CMV- or LTR-) regulated counterparts. We further show that RIP activity of both retrovectors mirrored fluctuations affecting endogenous INSULIN gene expression in human β cells. Finally, both α- and SIN γ-retrovectors were extremely poorly mobilized by the BXV1 xenotropic retrovirus, a common invader of human cells grown in immunodeficient mice, and, most notably, of human β cell lines. Conclusion Our novel α- and SIN γ-retrovectors are safe and convenient tools to stably and specifically express transgene(s) in mammalian β cells. Moreover, they both reproduce some regulatory patterns affecting INSULIN gene expression. Thus, they provide a helpful tool to both study the genetic control of β cell function and monitor changes in their differentiation status.http://link.springer.com/article/10.1186/s12896-019-0531-9α-RetrovectorSIN γ-retrovectorPancreatic β cell linesEndoC-β H2 cellsRat insulin promoterBXV1 xenotropic retrovirus |
| spellingShingle | Olivier Albagli Alicia Maugein Lukas Huijbregts Delphine Bredel Géraldine Carlier Patrick Martin Raphaël Scharfmann New α- and SIN γ-retrovectors for safe transduction and specific transgene expression in pancreatic β cell lines BMC Biotechnology α-Retrovector SIN γ-retrovector Pancreatic β cell lines EndoC-β H2 cells Rat insulin promoter BXV1 xenotropic retrovirus |
| title | New α- and SIN γ-retrovectors for safe transduction and specific transgene expression in pancreatic β cell lines |
| title_full | New α- and SIN γ-retrovectors for safe transduction and specific transgene expression in pancreatic β cell lines |
| title_fullStr | New α- and SIN γ-retrovectors for safe transduction and specific transgene expression in pancreatic β cell lines |
| title_full_unstemmed | New α- and SIN γ-retrovectors for safe transduction and specific transgene expression in pancreatic β cell lines |
| title_short | New α- and SIN γ-retrovectors for safe transduction and specific transgene expression in pancreatic β cell lines |
| title_sort | new α and sin γ retrovectors for safe transduction and specific transgene expression in pancreatic β cell lines |
| topic | α-Retrovector SIN γ-retrovector Pancreatic β cell lines EndoC-β H2 cells Rat insulin promoter BXV1 xenotropic retrovirus |
| url | http://link.springer.com/article/10.1186/s12896-019-0531-9 |
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