Fig 1: Cellular distribution of ARPC5 isoforms.(A) Shown are representative spinning-disk confocal images (maximum projection) of ARPC5.mCherry and ARPC5L.mCherry distribution in unstimulated JNLA cells. White arrows point to the respective C5 or C5L punctae seen in the nucleus. Also see related Figure 5—figure supplement 1A and B. (B) Subcellular distribution of the ARPC5 subunit and its isoform ARPC5L was determined by biochemical fractionation of JNLA cells with respective knockout of either C5 or C5L in the bulk culture. Representative immunoblots reveal levels of ARPC5 isoforms in the whole cell extract (WCE), cytoplasmic (C), and nuclear (N) fractions in the indicated JNLA knockout cells post fractionation. GAPDH and hnRNPL were used as markers for cytoplasmic and nuclear compartments, respectively. (C) Representative, deconvoluted, and segmented stimulated emission depletion (STED) single-plane images show endogenous nuclear actin filaments (stained with Phalloidin-647N) and ARPC5.mCherry/ARPC5L.mCherry (signal enhanced with anti-mCherry with secondary antibody in atto-594 channel) in A3.01 T cells, stimulated with PMA/ionomycin (P/I) for 30 s. Arrows (in white) point to the colocalization events. Percent colocalization is mentioned as mean ± SD (in white bar, top right) for each of the isoforms from three independent experiments. Scale bar, 500 nm. (D) The dot plot shows the frequency of colocalization of ARPC5 and ARPC5L with nuclear F-actin (from representative STED-deconvolved and segmented super-resolved images shown in Figure 5C) in A3.01 cells post 30 s of stimulation with PMA + ionomycin. Each dot represents colocalization events per cell that was analyzed. Source data is avaialble at https://doi.org/10.11588/data/YVYEO8.