Extended Data Fig. 2: Overexpression of single hFWE isoforms in MCF-7 hFWEKO cells does not alter viability.
a, MCF-7 hFWEKO cells were infected with each hFWE1/2/3/4–GFP/RFP construct independently and the cells were monitored via live-cell imaging. Images at 24 h show that MCF-7 hFWEKO cells homogenously expressing single hFWE isoforms do not undergo cell death (n = 3 biologically independent experiments, P values shown, two-tailed t-test). b, Analysis of live-cell imaging is shown as heat maps to represent the total number of cells, number of cells dying every 1.5 h, average cellular volume and average cellular sphericity; each block represents a gradient scale of low (blue), medium (yellow) and high (red) number or shape or size of the cells (n = 3, analysis performed using manual tool from Fiji and automated tools from Imaris, Genie tool used for representing data as heat maps). c, qPCR analysis of MCF-7 hFWEKO cells expressing each of four hFWE isoforms showed comparable lentiviral-mediated expression of hFWE1, hFWE2, hFWE3 or hFWE4 (n = 3 biologically independent experiments, mean ± s.d., ANOVA showed no significant differences). d, Flow cytometry-based annexin-V staining shows that overexpression of individual hFWE1/2/3/4–GFP/RFP isoforms does not induce apoptosis in MCF-7 hFWEKO cells (n = 3 biologically independent experiments, mean ± s.d., ANOVA showed no significant differences). e, Annexin-V staining and flow cytometry-based cell imaging (image stream) also show that overexpression of individual hFWE isoforms does not induce apoptosis (n = 3). f, Long-term effect of overexpression of single hFWE isoforms with GFP or RFP reporters over 21 days using clonogenic assay. The overexpression of hFWE isoforms does not affect the colony formation ability of MCF-7 hFWEKO cells (n = 3).
