The role of Diacylglycerol o-acyltransferases 1&2 (DGAT1 & DGAT2) in obesity mediated colon cancer growth
Colon cancer is the 3rd most diagnosed cancer in the United States and individuals that are obese have increased colon cancer risk, progression, reoccurrence, and resistance to therapy. Here, we aim to determine the role of two diacylglycerol O-acyltransferases, DGAT1 and DGAT2, in obesity mediated colon cancer growth. DGAT1 and DGAT2 (DGATs) are enzymes responsible for the initiation and growth of lipid droplets (LDs), intracellular organelles that store lipids. Limited findings show that increase LDs accumulation in colonic cells might be one of the mechanisms how obesity facilitates colon cancer growth. Colon cancer samples obtained from local patients show an increase in both DGATs levels in tumor tissues compared to uninvolved and this increase is further augmented in obese patients. In addition, DGATs levels are increased in human colon cancer cell lines (vs normal), and further augmented by obesity mediators. We found under oleic acid (OA) stimulation, increase in DGAT2 expression is regulated by myc binding to its promoter. Additionally, OA leads to systemic myc regulation of genes involved with cell growth and proliferation, tumor morphology, cancer, and gastrointestinal diseases (CUT&RUN). Further, we found a link between OA stimulated loss of FOXO3 and subsequent activation of pathways by myc. Inhibition of DGAT1 in human colon cancer cells leads to enlargement of existing LDs in the cytoplasm, while with DGAT2 inhibition LDs are trapped in ER. This suggests the important role of DGAT1 and DGAT2 in not only initiation and growth of LDs, but in LD trafficking in cells. Furthermore, DGATs inhibition attenuates colony formation of colonic cancer cells, and BrdU incorporation in newly synthesized DNA. This DGATs inhibition leads to cell cycle arrest at G1/S cell cycle, and recovers OA mediated loss of FOXO3, leading to FOXO3 presence on the promoter region p27kip1, a cell cycle inhibitor. In addition, we determined systemic transcriptional changes in colonic cells when DGATs are inhibited relative to untreated cells (RNAseq). We found over 7000 differentially expressed genes (DEG) related to cancer, gastrointestinal diseases, cellular growth and proliferation, cellular movement, cell cycle, immune response, and lipid metabolism. When molecular pathways related to these DEGs were compared to colon of high-fat diet-obese mouse, we found an opposite trend in regulation suggesting DGATs inhibition targets pathways driven by high fat diet. Similarly, opposite trends were seen in pathways when DGATs inhibited DEGs were compared to transcriptome from human colon cancer. We then validated expression of select genes using two colon cancer cell lines, HT29 and HCT116, treated with DGATs inhibitors. Several of these DEGs show an opposite response to DGATs inhibition between the HT29 and HCT116 cell lines. As HT29 cells have p53 mutation, we determined if these DGATs mediated differences between cell lines are dependent on p53. Using HCT116 p53 null cell (vs wildtype), we found that for two genes DGATs mediated reduction in expression is in part p53 dependent. Together, we demonstrated the significance of increased DGAT1 and DGAT2, responsible for elevated lipid droplet accumulation, in obesity mediated in colon cancer progression especially when driven by obesity.