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. 2018 Sep 28;8:14551. doi: 10.1038/s41598-018-32785-5

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© The Author(s) 2018

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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IFITMs inhibit HIV viral output and viral protein production in transfected cells. HEK293T cells were transfected with a titration of expression vectors for FLAG tagged IFITMs, with the total quantity of 0.5 μg, plus 0.5 μg of HIV-1 NL4-3 proviral DNA. (A) Levels of virus production were measured by p24 ELISA 48 h post-transfection, and (B) intracellular viral proteins and IFITMs were analyzed by immunoblotting. HEK293T cells were transfected with 0.5 μg of FLAG-tagged IFITM expression vectors and 0.5 μg of HIV-1 NL4-3 proviral DNA. Levels of virus production from indicated HIV-1 proviral DNAs were measured by (C) p24 ELISA and (D) reverse transcriptase (RT) activity assay and (E) immunoblotting of intracellular proteins 48 h post-transfection. (F) Mean fluorescence intensity (MFI) of GFP expression in HEK293T cells co-transfected with 0.5 μg CMV-driven GFP vector and 0.5 μg IFITM-expression vectors or empty vectors measured by flow cytometry 48 hours post-transfection. (G) HIV-1 NL4-3 and 89.6 viruses were produced from HEK293T cells transfected with vector or the indicated IFITMs in TZM-bl cells for 48 hours. Infectivity in TZM-bl cells was measured by luciferase activity assay 48 hours after infection with virus stock with equivalent p24 concentration. Data show mean + S.E.M. of more than 3 independent experiments. All differences were assessed with Student’s t-test and *indicates p < 0.05.