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. 2018 Feb 16;8:3217. doi: 10.1038/s41598-018-21344-7

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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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Heparan sulfate expression is missing on ITK deficient cells but is not responsible for reduced viral binding. (A) Jurkat cells expressing ITK (no shRNA, n.t. shRNA) or not expressing ITK (258 shRNA, 614 shRNA) were examined by FACS analysis for heparan sulfate on the cell surface using anti heparin sulfate-FITC. (B) Jurkat wild-type cells were treated with a mixture of 5 units (U) heparinase I and III and analyzed by FACS again for heparan sulfate expression using anti heparin sulfate-FITC, proofing the efficiency of enzymatic digestion. Filled histograms represent the isotype control. (C) Jurkat wild-type cells with enzymatically removed heparan sulfate (5 U heparinase I and III mixture) tested for binding of GFP-labelled HIV-1 virus particles pseudotyped with HIV-1 envelope protein. (D) Jurkat cells expressing ITK (no shRNA, n.t. shRNA) or not expressing ITK (258 shRNA, 614 shRNA) were analyzed by FACS analysis for CD47 using anti-CD47-FITC antibody. Filled histograms represent the isotype control. All figures show one representative experiment out of three repetitions.