Journal: Journal of cell science
Article Title: A role for class I p21-activated kinases in the regulation of the excitability of the actin cytoskeleton.
doi: 10.1242/jcs.263763
Figure Lengend Snippet: Fig. 4. Formation of cortactin/actin rings is PI3K dependent. (A) Still from Movie 6 of an MEF transiently overexpressing mCherry–LifeAct and PI(3,4,5)P3 marker (GFP BTK PH) following treatment with 10 μM G5555 for at least an hour. Accompanying linescan demonstrates localisation PI(3,4,5)P3 marker relative to actin cytoskeleton. (B) Still from Movie 7 of an MEF transiently overexpressing mEGFP–LifeAct and PI(4,5)P2 marker (mCherry PH domain of PLCdelta1) following treatment with 10 μM G5555 for at least an hour. Accompanying linescan demonstrates localisation of PI(4,5)P2 marker relative to actin cytoskeleton. (C) Percentage of cells producing cortactin rings in the presence and absence of 10 μM PI3K inhibitor LY294002 alongside either cotreatment with 10 μM G5555 or overexpression of GFP–PAK1Δkin. At least 100 cells analysed per condition for each of at least three biological repeats. Inhibitors were added 1 h before fixation. (D) Schematic summarising the localisation of various proteins upon addition of G5555. Scale bars: 10 μm. All error bars indicate s.d. ***P≤0.001 (ordinary one-way ANOVA followed by a post hoc Tukey’s multiple comparison test). A.U., arbitrary units.
Article Snippet: All other plasmids were acquired from Addgene: mApple LifeAct (Addgene plasmid #54747), mEGFP LifeAct (Addgene plasmid #54610), mCherry N-WASp (Addgene plasmid #55164), Myo1e mCherry (Addgene plasmid #27698), EmeraldVinculin (Addgene plasmid #54303), mCherry Arp2 (Addgene plasmid #54980), GFP BTK PH (Addgene plasmid #51463), mCherry PH domain of PLCdelta1 (Addgene plasmid #36075), GFP-PH-Tapp1 (Addgene plasmid #161985), p40PX-EYFP (Addgene plasmid #19011), Cyfip GFP (Addgene plasmid #109139).
Techniques: Marker, Over Expression, Comparison
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