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. 2019 May 1;9:6776. doi: 10.1038/s41598-019-43255-x

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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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PFBC XPR1/SLC53A1 variants and XPR1, PiT1 and PiT2 cell surface expression. (a) Cell surface detection of XPR1/SLC53A1, PiT1/SLC20A1 and PiT2/SLC20A2 with RBD specific ligands derived from mouse xenotropic MLV, koala endogenous retrovirus, and mouse amphotropic-MLV, respectively (open histograms); HEK293T positive and negative control cells for XPR1/SLC53A1 expression were transfected with either an anti-luciferase siRNA (siLUC) or an anti-XPR1/SLC53A1 siRNA (siXPR1), respectively. WT XPR1/SLC53A1 and variants were assayed for cell surface expression upon co-transfection of siXPR1 alone (−) or in combination with an expression vector coding either for the HA-tagged WT XPR1 (WT), or the PFBC XPR1 C-terminal variants (R459C, N619D, or I629S), or the p.(L612_696Tdel) artificial XPR1 mutant (Δ612–696). Numbers indicate the delta mean fluorescence intensity of a representative experiment (n = 3) compared to the non-specific staining obtained with the secondary IgG antibody (grey histograms). (b) Representative immunoblot of HA-tagged XPR1/SLC53A1-containing cell lysates with an anti-HA antibody (upper panel), or with an anti β-actin antibody used as loading control (lower panel).