Objective:To investigate the mechanism of action and prognostic value of Dynamin 3 (DNM3) in gastric cancer.Methods:The bioinformatic analysis, experimental study and retrospective cohort study was conducted. The clinicopathological data, fresh gastric cancer tissues, paired normal tissues and the corresponding paraffin sections of 153 gastric cancer patients who underwent radical gastrectomy in Fujian Medical University Union Hospital from January 2013 to July 2018 were collected. Tissues and the corresponding paraffin sections were subjected to quanti-tative real-time polymerase chain reaction, immunoblotting assay, flow cytometric cell cycle assay and immunohistochemical staining, respectively, and clinicopathological data were used for prognostic analysis. The stomach adenocarcinoma (STAD) dataset from the Cancer Genome Atlas (TCGA) database was collected for bioinformatic analysis. Observation indicators: (1) DNM3 gene expression in TCGA-STAD in gastric cancer; (2) mutations and copy number alterations of DNM3 in gastric cancer; (3) methylation level of promoter of DNM3 in gastric cancer; (4) relative protein expression of DNM3 and p53 in gastric cancer; (5) DNM3 correlation and enrichment analysis; (6) ratio of G0/G1 phase, S phase and G2/M phase of cell cycle progression; (7) correlation between immune cell infiltration and DNM3 in gastric cancer; (8) correlation between results of immunohistochemical (IHC) staining and clinical features; (9) analysis of independent factors influencing 5-year overall survival rate of gastric cancer patients. Measurement data with normal distribution were represented as Mean±SD, and comparison among multiple groups was conducted using the ANOVA and further comparison between two groups was conducted using the LSD. Comparison between two groups was conducted using the t test. Measurement data with skewed distribution were represented as M( Q1, Q3), and comparison between groups was conducted using the Mann-Whitney U test. Count data were described as absolute numbers or percentages, and compari-son between groups was conducted using the chi-square test or Fisher exact probability. Comparison of ordinal data was conducted using the rank sum test. The Pearson correlation coefficient or Spearman correlation coefficient was used to test the correlation between groups. Univariate and multivariate analyses were conducted using the COX proportional risk regression model. The Kaplan-Meier method was used to draw survival curves and calculate survival rates, and the Log-Rank test was used for survival analysis. The Benjamini-Hochberg false discovery rate correction was used for adjusting of the P-value. Results:(1) DNM3 gene expression in TCGA-STAD. The expression levels of DNM3 gene in the 27 tumor tissues and paired normal tissues of the TCGA-STAD database were 0.775(0.605,1.161) and 1.216(0.772,1.681), showing a significant difference between them ( Z=?2.64, P<0.05). The messenger RNA (mRNA) expression levels of DNM3 gene in 48 pairs of gastric cancer tissues and paired normal tissues of the author′s center were 4.370(2.870,6.040) and 2.520(0.850,4.170), showing a significant difference between them ( Z=?4.39, P<0.05). (2) Mutations and copy number alterations of DNM3 in gastric cancer. There were 16 gastric cancer patients in the TCGA-STAD database with DNM3 mutation or somatic copy number alterations, including 6 cases with missense mutations, 1 case with truncated mutation, 8 cases with copy number gain and 1 case with copy number loss. The mRNA expression levels of DNM3 gene before and after mutation in the 370 gastric cancer patients of the TCGA-STAD database were 6.13(5.40,7.08) and 5.02(3.98,5.46), showing a significant difference between them (Log 2FC=?1.11, Z=?2.59, P<0.05). (3) Methylation level of promoter of DNM3 in gastric cancer. There were 372 gastric cancer patients in the TCGA-STAD database undergoing DNM3 methylation and mRNA examinations, and the results showed that levels of methylation and mRNA expression of DNM3 was 0.198 (-0.458, 0.301) and 6.014 (5.141, 6.628), respectively. The levels of methylation in DNM3 was negatively correlated with its mRNA expression ( r=?0.38, P<0.05). Results of follow-up in 32 patients showed that the 3-year overall survival rate of 16 cases with high levels of methylation in DNM3 and 16 cases with low levels of methylation in DNM3 was 18.8% and 41.3%, respectively, showing a significant difference between them ( hazard ratio=1.40, P<0.05). Results of immunoblot-ting assay showed that the relative expression level of DNM3 protein in the AGS cells treated with 0, 0.5, and 1.0 μmol/L of 5-azacytidin was 0.270±0.020, 0.357±0.051 and 0.599±0.039, respectively, showing a significant difference among the three groups ( F=57.84, P<0.05). The relative expression level of DNM3 protein in the HGC-27 cells treated with 0, 0.5, and 1.0 μmol/L of 5-azacytidin was 0.316±0.038, 0.770±0.031 and 0.877±0.052, respectively, showing a significant difference among the three groups ( F=156.30, P<0.05). (4) Relative protein expression of DNM3 and p53 in gastric cancer. Results of immunoblotting assay showed that the relative expression of DNM3 and p53 protein was 0.688±0.047 and 0.872±0.041 in the AGS cells transfected with pCMV-DNM3 plasmid, versus 0.249±0.029 and 0.352±0.020 in the AGS cells transfected with control plasmid, showing significant differences in the above indicators between the two types of cells ( t=13.77,19.74, P<0.05). The relative expression of DNM3 and p53 protein was 0.969±0.069 and 1.464±0.081 in the HGC-27 cells transfected with pCMV-DNM3 plasmid, versus 0.456±0.048 and 0.794±0.052 in the HGC-27 cells transfected with control plasmid, showing significant differences in the above indicators between the two types of cells ( t=10.57, 12.06, P<0.05). (5) DNM3 correlation and enrichment analysis. Results of correlation analysis showed that DNM3 was positively correlated with genes such as RBMS3, CNTN4 and PDE1A ( r=0.52, 0.52, 0.50, P<0.05) and negatively correlated with genes such as SLC25A39, PAICS and GAPDH ( r=?0.41, ?0.40, ?0.40, P<0.05) in gastric cancer. Results of gene set enrichment analysis showed that the set of genes related to ribosome and oxidative phosphorylation were upregulated in gastric cancer patients with DNM3 low expression [normalized enrichment score (NES)=?3.30, ?2.16, P<0.05], while the set of genes related to immunomodulatory interactions between lymphocytes and non-lymphoid cells were upregulated in gastric cancer patients with DNM3 high expression (NES=1.67, P<0.05). Results of gene ontology analysis showed that the low expression of DNM3 was associated with the separation of mitotic sister chromatid (No.0000070), nonsense-mediation of nuclear transcriptional mRNA catabolic process, sister chromatid separation (No.0000819), nuclear transcriptional mRNA catabolic process and regulation of oxidative phos-phorylation (NES=?2.29, ?3.10, ?2.33, ?2.56, ?2.68, P<0.05). Results of Kyoto encycl opedia of genes and genomes analysis showed that metabolic pathway related to ribosome and oxidative phosphory-lation were upregulated and crosstalked in gastric cancer with low expression of DNM3 (NES=?3.34, ?2.21, P<0.05). (6) Ratio of G0/G1 phase, S phase and G2/M phase of cell cycle progression. Results of flow cytometric cell cycle experiments showed that the proportions of G0/G1 phase, S phase and G2/M phase in the cell cycle was 65.1%±3.0%, 17.3%±3.0% and 17.6%±1.0% in the AGS cells transfected with pCMV-DNM3 plasmid, versus 53.4%±4.0%, 26.3%±2.0% and 20.3%±3.0% in the AGS cells transfected with control plasmid, showing significant differences in the proportions of G0/G1 phase and S phase in the two types of cells ( t=4.05, 4.32, P<0.05). (7) Correlation between immune cell infiltration and DNM3 in gastric cancer. Results of immune cell infiltration examination showed that the expression level of DNM3 was positively associated with mast cells, NK cells, pDCs, B cells, follicular helper T cells, effector memory T cells, T cells, central memory T cells, CD8 T cells, DC cells, macrophages, γ-δ T cells (Tgd), iDCs and eosinophils infiltration (Spearman correlation coefficients as 0.41, 0.29, 0.26, 0.20, 0.22, 0.22, 0.13, 0.16, 0.15, 0.14, 0.14, 0.17, 0.18, 0.22, P<0.05) and negatively associated with Th17 cell, Th2 cells and NK CD56 dim cells infiltration ( r=?0.18, ?0.23, ?0.10, P<0.05). (8) Correlation between results of IHC staining and clinical features. Results of IHC staining analysis showed that the IHC score of DNM3 was 3(2,4) in the 105 gastric cancer tissues, versus 6(4,9) in the 105 paired normal tissues, showing a significant difference between them ( Z=-7.35, P<0.05). There were significant differences in gender, tumor location and N stating between the 70 patients with low expression of DNM3 and the 35 patients with high expression of DNM3 ( χ2=4.29, 7.67, 6.86, P<0.05). (9) Analysis of independent factors influencing 5-year overall survival rate of gastric cancer patients. Results of multivariate analysis showed that stage pT3?4 and low IHC score of DNM3 were independent risk factors for 5-year overall survival rate of gastric cancer patients ( hazard ratio=1.91, 0.51, 95% confidence interval as 1.06?3.43, 0.26?0.98, P<0.05). The 5-year overall survival rate was 44.3% in patients with low expression of DNM3, versus 65.7% in gastric cancer patients with high expression of DNM3, showing a significant difference between them ( χ2=5.02, P<0.05). Conclusion:DNM3 is a tumor suppressor and an independent predictor of poor prognosis for gastric cancer, which may regulate gastric cancer cell cycle and immunosuppression in the tumor microenvironment through methylation.