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. 2017 Jul 12;7:5209. doi: 10.1038/s41598-017-05487-7

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© The Author(s) 2017

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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TRIM21 mediates Lys63-linked ubiquitination around type II Toxoplasma. (a) Representative immunofluorescence confocal images of IFNγ-stimulated wild-type (upper panel) or TRIM21 knockout (lower panel) mouse embryonic fibroblasts infected 1 h with type II Toxoplasma expressing GFP and stained for Lys48 ubiquitin (left panel) or Lys63 ubiquitin (right panel). Scale bar is 5 μm. (b) Quantification of ubiquitin- (Ub), Lys48 ubiquitin- and Lys63 ubiquitin-positive parasite-containing vacuoles in IFNγ-stimulated wild-type versus TRIM21 knockout MEFs infected 1 h with type II Toxoplasma. Data were pooled from three independent experiments. Mean ± SEM, *p < 0.05, **p < 0.005, 2-way ANOVA. (c) Quantification of Lys63 ubiquitin-positive parasite-containing vacuoles in IFNγ-stimulated mouse embryonic fibroblasts infected 1 h with the canonical Toxoplasma type III (CEP) strain and Toxoplasma type III strain transgenic for the type I version of Rop18 (CEP Rop18I). Mean ± SEM, **p < 0.005, 2-way ANOVA. (d) Quantification of GBP1 and GBP2 localisation to the parasite-containing vacuoles in IFNγ-stimulated wild-type versus TRIM21 knockout mouse embryonic fibroblasts 1 h p.i. Data pooled from three independent experiments, unpaired t-test.