Fig. 5: Screening for effectors of YAP1 for survival of an ALC-resistant cell line. | Nature Communications

Fig. 5: Screening for effectors of YAP1 for survival of an ALC-resistant cell line.

From: YAP1 mediates survival of ALK-rearranged lung cancer cells treated with alectinib via pro-apoptotic protein regulation

Fig. 5

a Schematic explanation for the establishment of H2228-ARY. This YAP1-activated cell line was established by exposing NCI-H2228 cells to ALC for 3 months and subsequent cell cloning. b Representative images of H2228-ARY with a 96-h exposure to either ALC or vehicle. Fixed cells were stained by YAP1-alexa488 and Hoechst. Scale bar = 100 µm. c Quantified nuclear localization of YAP1 in H2228 or H2228-ARY cells. The number of images analyzed is presented in the figure. Significance was evaluated with a one-way ANOVA followed by Sidak’s test. d A cell viability assay when H2228 parent and H2228-ARY cells were exposed to stepwise concentrations of ALC for 96 h. Significance was evaluated using a two-way ANOVA followed by Dunnett’s test comparing viability with that of H2228 parent cells. e A cell viability assay when H2228-ARY cells were treated with ALC under the genetic inhibition of YAP1. Significance was evaluated using a two-way ANOVA followed by Dunnett’s test comparing viability with that of siNC-treated cells. f An apoptosis assay on H2228-ARY cells treated with ALC under siYAP1. Significance was evaluated using a one-way ANOVA followed by Sidak’s test. g Screening of effectors for YAP1 that induced apoptosis using qRT-PCR. Heatmap of the 13 factors (left, n = 4 independent experiments) and detected individual values of MCL1 and BCLXL gene expression (right, n = 8 independent experiments). The individual values of other genes were plotted in Supplementary Fig. 5a. Color scale shows the change of each gene expression: magenta and green means increased and decreased expression, respectively. Significance was evaluated using a one-way ANOVA followed by Sidak’s test. h The knockdown of YAP1 inhibited the protein expression of MCL1 and BCLXL in H2228-ARY cells. i Summary of exome sequencing that compared the sequences of presented genes between H2228 cells and H2228-ARY cells. The full results are shown in Supplementary Data 3. The exome sequence can identify some parts of the intron sequence that are near the exons. ALC alectinib, VER:verteporfin, siSC si scramble control. Error bars indicate ± S.D. *P < 0.05.

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