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. 2015 Jan 1;26(1):55–65. doi: 10.1091/mbc.E14-07-1200

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© 2015 Havrylenko, Noguera, et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

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FIGURE 1: — WAVE binds Ena/VASP for increased motility in vitro. (a) Scheme of general Ena/VASP and WAVE domain organization, with the putative interaction between the two marked by a double arrow. (b) Immunolabeling of beads coated with different PRD-VCA and VCA constructs incubated in either full-length VASP or ΔEVH1-VASP (lacking the putative site for interaction with WAVE). Only beads carrying the PRD domain light up and only when incubated in VASP possessing its EVH1 domain; p < 0.0001. PRD-VCA-WAVE and PRD-VCA-WASP beads in VASP are also significantly higher than VCA in VASP, p < 0.0001, not marked on the graph for clarity. Left, fluorescence intensity measurements; right, representative images. From 20 to 50 beads were analyzed per condition. Epifluorescence microscopy. (c) Comets on PRD-VCA-WAVE beads in the presence of wild-type VASP and ΔEVH1-VASP and with no addition. Actin comets appear as darker streaks behind the beads, which appear white. All pictures were taken at ∼10- to 15-min reaction time. In the graph, speeds for PRD-VCA-WAVE beads are represented normalized to wild-type VASP addition to account for day-to-day variations. No addition and addition of ΔEVH1-VASP give speeds that are 60% that of wild type, p = 0.004 and 0.003, respectively. PRD-VCA-WAVE beads moved at speeds of 0.3–1.4 μm/min, depending on the day and the additive. Phase contrast microscopy. All data are represented as averages ± SD. p values calculated with the Student's t test. Bars, 1 μm.