The Role And Molecular Mechanism Of P2 Promoter-driven Hepatocyte Nuclear Factor-4? In The Migration And Invasion Of Gastric Cancer

Hepatocyte nuclear factor 4 alpha(HNF4a),a highly conserved member of the nuclear receptor superfamily of transcription factors,commonly known as the master regulator of hepatic proliferation,differentiation,metabolism.HNF4a can be regulated by two promoters,P1 and P2(named P1-HNF4a and P2-HNF4a,respectively),and their expression,tissue distribution and function are quite different.Whereas the role of HNF4a in the regulation of liver and intestines is well recognized and extensively studied,its expression and function in gastric cancer especially in advanced and recurrent metastatic gastric cancer are still poorly understood.In our study,we found that P2 promoter-driven hepatocyte nuclear factor 4 alpha(P2-HNF4a)was highly expressed in the recurrence and metastasis of gastric cancer and the expression in the metastatic lesion higher than that in the original lesion,suggesting that it may be related to the migration and invasion of gastric cancer.We found that knockdown of P2-HNF4a or treatment with HNF4a antagonist B16015 could significantly inhibit the migration ability in gastric cancer cells AGS by live cell movement trajectory,3D spheroid cell migration and invasion experiment,Wound healing and Transwell experiment.Conversely,overexpression of HNF4a7(one HNF4a subtype of P2 promoter-driven expression)significantly enhanced the migration and invasion ability in gastric cancer cells HGC-27.We also verified the same results in a nude mouse gastric cancer metastasis model by intraperitoneal injection;Furthermore,we found that the ability of P2-HNF4a to promote the invasion and migration in gastric cancer cells depends on its transcriptional activation activity.Then,by using cell-level transcriptome sequencing,metabolome analysis and their correlation analysis,we found that P2-HNF4a may play a role in promoting the migration and invasion of gastric cancer on one hand by regulating the expression of lectin-like oxidized low-density lipoprotein receptor-1(OLR1)and thereby activating the PI3K/Akt/GSK3? signaling pathway;On the other hand,they may upregulate the glycolytic pathway related enzymes(hexokinase(HK),etc)and the pentose phosphate pathway related enzymes(glucose-6-phosphate dehydrogenase(G6PD),etc)expression,and then remodel the energy metabolism of tumor cells,(namely promoting aerobic glycolysis and the pentose phosphate pathway,the Warburg effect),thereby promoting migration and invasion of gastric cancer cells.To sum up,for the first time,our present study revealed the role and the underlying molecular mechanisms of P2-HNF4a in promoting the migration and invasion of gastric cancer,namely regulating the expression of OLR1 and glycolytic pathway related enzymes and the pentose phosphate pathway related enzymes expression,then remodel the energy metabolism of tumor cells through its transcriptional activation activity,thereby promoting the migration and invasion in gastric cancer.