cGAS and lamin A localization at aggregate induced rupture sites. (A) Representative image of U2OSWT cells expressing polyQ74-NLS (green), immunostained for endogenous cGAS (magenta). Profile plot of line scan in inset, showing localization of endogenous cGAS around a nuclear aggregate. (B) Representative stills from live-cell imaging of U2OSWT cells expressing polyQ74-NLS (green) and mCherry-cGAS (magenta). Time-lapse zooms show multiple ruptures (Rupture 1, 2 and 3) occurring at different aggregates. (C) Graphs of intensity profiles across rupture sites shown in B, indicating mCherry-cGAS accumulation around individual aggregates. (D) RPE-1 cells expressing polyQ74-NLS aggregates and mCherry-cGAS. Orange arrows indicate nuclear cGAS accumulation at polyQ aggregates. (E and G) Representative U2OS-RFP-NLS cells expressing polyQ74-NLS (green) and HaloTag-lamin A (magenta). Zooms of E showing aggregate growth during imaging. (F) Timelapse of zooms of cell shown in E. Stills show HaloTag-lamin A scar formation upon rupture (black arrows) at nuclear polyQ74-NLS aggregates. (H) Time-lapse images of the cell shown in G with minor HaloTag-lamin A accumulation (black arrow) at a nuclear aggregate. PolyQ and cGAS signal was gamma adjusted (γ = 0.75). Scale bars represent 10 μm (A, B, D, E, and G) and 5 μm (Azoom, Ezoom, F, and H).
