Supplementary Figures 1 through 11 from Phospholipase D1 Acts through Akt/TopBP1 and RB1 to Regulate the E2F1-Dependent Apoptotic Program in Cancer Cells
Supplementary Figure S1. PLD1 inactivation promotes apoptosis under serum deprived condition. Supplementary Figure S2. Scatter diagrams of gene expression microarray by depletion of PLD1. Supplementary Figure S3. PLD1 inactivation decreases RB1 expression in the tumor tissues from ApcMin/+ mice. Supplementary Figure S4. Loss of PLD1 decreases RB1 expression in the tumor tissues from AOM/DSS mice. Supplementary Figure S5. Epigenetic regulation of RB1 promoter is not affected by depletion and inhibition of PLD1. Supplementary Figure S6. Pre-miR-4465 and -192 decrease RB1 mRNA level. Supplementary Figure S7. IEC-specific PLD1 overexpression in ApcMin/+ mice increases the levels of RB1 and p-Akt. Supplementary Figure S8. PLD1 inhibition decreases Akt phosphorylation in AOM/DSS mice. Supplementary Figure S9. Effect of depletion of p27KIP1, Bim, or NOXA on E2F1-induced apoptosis. Supplementary Figure S10. Effect of RB1, Akt, and anti-miR-4496/-192 on PLD1 inhibition-repressed beta-catenin and c-Myc expression. Supplementary Figure S11. Multivariate survival analysis between expression of PLD1 and proapoptotic E2F1 target genes in CRC patients.
Funding
National Research Foundation of Korea
Translational Research Center for Protein Function Control
Ministry for Health, Welfare, and Family Affairs
Korean Health Technology R&D Project
History
ARTICLE ABSTRACT
The RB1/E2F1 signaling pathway is frequently deregulated in colorectal cancer and has been suggested to intersect with Wnt/β-catenin and PI3K/Akt pathways, but molecular evidence for this link is lacking. In this study, we demonstrate that phospholipase D1 (PLD1), a transcriptional target of β-catenin/TCF4, orchestrates functional interactions between these pathways during intestinal tumor development. Overexpression of PLD1 in intestinal epithelial cells protected cells from apoptosis induced by PLD1 ablation in the Apcmin/+ mouse model of intestinal tumorigenesis. Mechanistic investigations revealed that genetic and pharmacologic targeting of PLD1 promote the E2F1-dependent apoptotic program via both miR-192/4465–mediated downregulation of RB1 and inhibition of Akt–TopBP1 pathways. Moreover, the miRNA–RB1 axis and Akt pathway also contributed to the PLD1-mediated self-renewal capacity of colon cancer–initiating cells. Finally, PLD1-driven E2F1 target gene expression positively correlated with tumor stage in patients with colorectal cancer. Overall, our findings suggest that PLD1 mediates cross-talk between multiple major signaling pathways to promote the survival and malignancy of colon cancer cells and may therefore represent an ideal signaling node for therapeutic targeting. Cancer Res; 77(1); 142–52. ©2016 AACR.Usage metrics
