Dominant negative nesprin and SUN constructs displace endogenous nesprins from the nuclear envelope. A and B, immunofluorescence images of MEFs stably expressing DN KASH (A) or mCherry (B). Cells were stained for nesprin-3 (second panel) and DNA (Hoechst 33342, third panel). C and D, immunofluorescence images of MEFs transiently expressing DN SUN1L (C) or GFP alone (D). Cells were stained for nesprin-3 (second panel) and DNA (third panel). Localization of DN SUN1L to the nuclear envelope and endoplasmic reticulum is shown in supplemental Fig. S1. Scale bars, 10 μm. E, scheme of LINC complex disruption by DN KASH and the displacement of endogenous nesprins from the nuclear envelope to the ER. INM, inner nuclear membrane; ONM, outer nuclear membrane. F and G, percentage of cells with normal nuclear envelope (NE) localization of endogenous nesprin-3 in DN KASH (F) and DN SUN1L (G)-expressing cells. More than 100 cells were analyzed for each sample; data are represented as mean ± S.E.; *, p < 0.05.