Studies On The Functions And Mechanisms Of NAIF1in Gastric Cancer Cell

As a common digestive tract cancer, the incidence of gastric cancer occupied the fourth place of all human cancers with a lethality rate of70%, which is much higher than other common human cancers. High morbidity and high mortality rate of gastric cancer has a serious impact on human health. Tumor invasion and metastasis are the main reasons for the bad prognosis of patients and lead to high lethality rate, therefore, researching the related molecules and pathways of invasion and metastasis in gastric cancer cell will not only help us with a better understanding of the molecular mechanisms, but also provide new markers and targets for gastric cancer diagnosis and therapy.NAIF1is a nuclear apoptosis-inducing factor. Previous findings showed that NAIF1was able to induce apoptosis in HeLa cells and MKN45cells through mitochondrial pathway, and these events involved the activation of Caspase9and Caspase3, in addition, NAIF1showed a different expression between human gastric cancer tissues and adjacent tissues. However, whether NAIF1was involved in other cell functions and pathways was still unknown.In this study, we focus on the functions and mechanisms of NAIF1in gastric cancer cells. A variety of experimental techniques were employed including confocal scanning, flow cytometry, transwell, real-time Q PCR, two-dimensional electrophoresis and immunoblotting.As a result, we did not found any expression of NAIF1at RNA or protein level in gastric cancer cell AGS, MKN45, SGC7901or one immortalized gastric mucosa cell line GES-1. When NAIF1was transfected into AGS and MKN45cell, it showed that NAIF1only distributed in the nucleus. Overexpression of NAIF1led to a G1-S arrest in MKN45cells and also decreased the cell migration ability in both AGS and MKN45, with a decrement of3times in AGS or2.8times in MKN45. Immunoblotting result suggested that NAIFl induced cell cycle arrest by down-regulating the expression of CyclinDl. Besides, NAIFl was able to reduce the activity of Erkl/2and JNK with or without stimulation by UV/serum, but had no effect on P38. In addition, NAIF1was able to reduce the expression level of JNK. As a downstream gene of MAPK pathway, the down-regulation of expression of MMP2and MMP9were also observed. Moreover, inhibition of p-FAK was detected in NAIFl-overexpressed cell. Two-dimensional electrophoresis was used to indicate that overexpression of NAIF1had a effect on the protein expression profile in MKN45cells, with up-regulation of5proteins including PSMC2、PSMD13、TXNRD1、TCP1-gamma and NDUSF1, and down-regulation of3protein,14-3-3epsilon, RNH1and APIAOBP. Westernblotting was introduced to vertify the result of Two-dimensional electrophoresis.By using genomics approaches, we also found that155genes showed a>2fold change after overexpressing NAIF1in cell, correlation analysis showed most of which were involed in immune response pathways. These proteomics and genomics data provide new ideas and directions for researching NAIF1in the future.This is the first time to discover the ability of NAIFl in cell migration and the first time to investigate the relationship between NAIF1and MAPK pathway, as well as CyclinDl and other downstream moleculars of MAPK pathways. This research is an innovation by revealing the functions and molecular mechanisms in which NAIFl was involved, have directive significance for understanding function and molecular mechanism of NAIF1in future, and provide a new target for the therapy of tumor invasion and metastasis.