Lack of LCBs causes genomic instability. (a) Representative immunofluorescence images of RPE-1 cells upon knockdown of scramble (Control), SPTLC1, or SPTLC2 cultured for 96 h, or treated with 5 µM myriocin or 10 µM fumonisin B1 in lipid-depleted medium for 48 h. Green, anti-alpha-tubulin; purple, Hoechst 33342 (DNA); and red, anti-lamin B1. Scale bar, 5 µm. Some images are also shown in Fig. S4 c. (b) Quantification of cells with micronuclei. Error bars represent the standard deviation of three biological replicates (each replicate, n > 100 cells). *P < 1e-4, one-way ANOVA. (c) Representative immunofluorescence image of RPE-1 nuclei showing NE blebs upon knockdown of SPTLC1 or SPTLC2. Scale bar, 5 µm. (d) Representative immunofluorescence image of RPE-1 cells with an anaphase bridge upon knockdown of SPTLC1 or SPTLC2. Scale bar, 5 µm. (e) Quantification of cells with nuclear blebs. Error bars represent the standard deviation of three biological replicates (each replicate, n > 100 cells). *P < 1e-4, one-way ANOVA. (f) Quantification of cells with anaphase bridge. Error bars represent the standard deviation of 3 biological replicates (each replicate, n > 100 cells). *P < 1e-4, one-way ANOVA. (g) Representative immunofluorescence images of RPE-1 cells 45 min after release from a G2 arrest. Green, anti-gamma-tubulin; blue, Hoechst 33342 (DNA); and red, anti-CDK5RAP2 (centrosome). Scale bar, 5 µm. (h and i) Quantification of cells with misaligned chromosomes (h) or multipolar spindle (i). Error bars represent the standard deviation of three biological replicates (each replicate, n > 100 cells). *P < 1e-3, one-way ANOVA.
