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. 2019 Feb 28;9:3134. doi: 10.1038/s41598-019-40003-z

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© The Author(s) 2019

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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Activated expression of endogenous BST-2 by lentiviral CRISPR transduction. (A) HOS cells were cotransduced with lentiviruses expressing dCas9-VP64, MS2-p65-HSF1, and either BST-2-targeting sgRNA (sgBST2) #1, #2, or the combination of #1 and #2 (sgBST2#1, sgBST2#2, or sgBST2#1/2, respectively). Cell extracts derived from transduced HOS cells were subjected to immunoblot analyses using an anti-BST-2 polyclonal antibody. β-actin was used as a loading control. (B) HOS cells transduced in A were cloned (designated HOS-sgBST2#x-x) and RNAs extracted from resultant cells were analyzed by real-time RT-PCR. Data were normalized to those of the housekeeping gene RPL27 mRNA and are shown as a fold difference in BST-2 copies compared with those in HeLa cells (mean ± s.d. from three independent experiments). (C,D) Control and cloned HOS cells together with HeLa cells were analyzed for cell-surface expression of BST-2 by flow cytometry (C) or for its intracellular expression by immunofluorescence (D; bars, 10 μm) using anti-BST-2 polyclonal antibodies.