The BRCA1 aggregates exclude large nuclear structures.

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The BRCA1 aggregates exclude large nuclear structures. The BRCA1 aggregates exclude large nuclear structures. (A) Accumulation of the proteasomal protein Pre6–GFP around BRCA1 aggregates. The depicted images result from live fluorescence microscopy analyses of Pre6–GFP cells expressing BRCA1 or BRCA1–mCherry. Pre6–GFP cells transformed with the vector plasmid are used as a negative control. Overlay images of GFP and mCherry signals (Merge) as well as transillumination images (Trans) are also shown. Arrows pinpoint the ‘hole’ pattern observed. The number of cells analyzed is indicated in Table S1. Scale bars: 2 μm. (B,C) 3D representation of fluorescence intensities measured in the squares drawn in A. The ‘hole’ pattern is present in the foreground of C. (D) Quantification of A (see Table S1 for details, including sample sizes). The star indicates a significant difference in proportions compared to vector cells (Fisher exact test). (E,F) Exclusion of the nuclear DNA from BRCA1 aggregates, using living Nup133–GFP cells expressing BRCA1–mCherry (E) or fixed wild-type cells expressing BRCA1–GFP (F). Cells with (top) or without (bottom) nuclear aggregate come from the same picture acquisition. Pictures are representative of 96 (E, top panel) and 8 (F, top panel) nuclear aggregates analyzed (100% exclusion). The nucleoporin Nup133–GFP allows the visualization of the nuclear membrane. DAPI stains the nuclear and mitochondrial DNA. Scale bars: 2 μm. Pierre Thouvenot et al. J Cell Sci 2016;129:4366-4378 © 2016. Published by The Company of Biologists Ltd