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. 2018 Apr 3;8:5503. doi: 10.1038/s41598-018-23816-2

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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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TAGLN2 inhibits Arp2/3 complex-mediated nucleation of branched filaments. (a) Time-based fluorometric analysis of pyrene-labelled actin polymerization. Polymerization assay in presence of His-TG2 (0, 2, 4, 8, 16, 32, and 64 μM) in F-buffer. Results are representative of at least three independent experiments. (b) Fitting of TG2 (red) and Arp2/3 (green) to F-actin reconstruction, showing the clash between TG2 and ARP2/3. (c) Dose-dependent inhibition of Arp2/3 and VCA-stimulated actin nucleation by TAGLN2. (0–32 μM TAGLN2 [TG2] or TAGLN2ΔAB, 20 nM Arp2/3, 200 nM GST_VCA). Results are representative of at least three independent experiments. (d) Competition with the Arp2/3 complex for actin binding in the presence of TAGLN2 or TG2ΔAB. Full-length blots/gels are presented in Supplementary Fig. S7. (e) Real-time imaging of actin branching formed by Arp2/3 and VCA with/without TAGLN2 (left). Scale bars, 2 μm. Quantification of branching density for each condition (right). *p < 0.05 versus no addition. (f) Fluorometric analysis of pyrene-labelled actin polymerization in the presence of the indicated proteins (Note, 2 μM actin, 8 μM TAGLN2 [TG2] or TAGLN2 mutants; 20 nM Arp2/3, and 200 nM GST_VCA). Results are representative of at least three independent experiments.