The Mechanism Of TMEM199 Promoting The Proliferation And Invasion Of Lung Adenocarcinoma Cells

Background: According to the latest cancer statistics released in 2020,lung cancer has become the most common type of cancer in the world and the leading cause of cancerrelated deaths.In the United States in 2020,there were approximately 228,820 new cases and 135,720 deaths.Lung cancer is mainly divided into two subtypes according to cell morphology,including non-small cell lung cancer(NSCLC)and small cell lung cancer.Lung adenocarcinoma(LUAD)and lung squamous cell carcinoma are the two main types of NSCLC.Among them,LUAD is the most common type,accounting for about 50% of lung cancer.For LUAD patients,although early diagnosis and treatment have improved and their survival results have been greatly improved,their 5-year survival rate is still unsatisfactory.New immunotherapy methods including the application of immune checkpoint inhibitors have significantly improved the efficacy and prognosis of some advanced lung adenocarcinomas.However,clinical predictors related to the prognosis of lung adenocarcinoma and immunotherapy are still lacking.TMEM199,a member of the transmembrane protein family,is the human homolog of the Candida albicans/yeast vacuolar ATPase(V-ATPase)assembly factor Vph2p/Vma12 p,which plays an indispensable key in the assembly process of the V-ATPase complex effect.At present,little is known about the role of TMEM199 in the human body,and its function in malignant tumors is rarely reported.At present,the role of TMEM199 in the occurrence and development of lung cancer is still unclear,and its underlying mechanism needs to be further clarified.Objective: To explore the expression and clinical significance of TMEM199 in lung cancer,and to explore its role and mechanism in the proliferation and invasion of lung adenocarcinoma cells.Meanwhile,to investigate the prognostic significance of TMEM199 expression in pan-cancer and its relationship with immune infiltration.Methods: The m RNA and protein expression levels of TMEM199 in human lung adenocarcinoma tissues were analyzed by bioinformatics,and the differential protein expression of TMEM199 was verified in 20 pairs of fresh lung cancer tissues.Western Blot,RT-q PCR,and immunofluorescence assays were used to detect the m RNA and protein expression levels of TMEM199 in lung adenocarcinoma cells as well as cell localization.The stable transfection technology was used to construct the stably expressed lung adenocarcinoma cell lines A549 and H1299 with overexpression and knockdown of TMEM199.MTT assay,Transwell invasion assay,and scratch assay were used to detect the effect of TMEM199 expression on the proliferation,invasion,and migration of lung adenocarcinoma cells.Western Blot was used to detect the effect of TMEM199 expression on the representative proteins of the Canonical Wnt signaling pathway and EMT process.The expression data,clinical survival data,and mutation data of 33 tumors were downloaded from the TCGA database.The expression and survival prognosis of TMEM199 in pan-cancer were analyzed by Limma package and Survival package in R language.The correlation between TMEM199 expression and immune microenvironment and immune infiltration in pan-cancer was investigated using Estimate,MCPCounter,and Cibersort algorithms.Limma package of R language was used to analyze the correlation between TMEM199 expression and immune checkpoint genes in pan-cancer.Perl programming language and R languages were used to summarize and analyze the correlation between TMEM199 expression and TMB and MSI in pan-cancer.Results: Compared with normal lung tissue,m RNA and protein expression levels of TMEM199 were significantly up-regulated in lung adenocarcinoma tissues,consistent results were obtained in lung adenocarcinoma cells.High expression of TMEM199 is closely associated with poor prognosis of OS and PFS in lung adenocarcinoma.Overexpression of TMEM199 can promote lung adenocarcinoma cells A549 and H1299 proliferation,invasion,and migration ability;The opposite result was obtained by knocking down the expression of TMEM199.Over-expression of TMEM199 significantly up-regulated the expression levels of active-β-catenin and c-myc in the Canonical Wnt signaling pathway of lung adenocarcinoma cells A549 and H1299.The expression of Ecadherin,an important EMT factor,was down-regulated,while the expression of vimentin was up-regulated.The opposite result was obtained by knocking down the expression of TMEM199.Pan-cancer bioinformatics analysis showed that TMEM199 was heterogeneous in 33 tumors,and the expression level of TMEM199 was generally upregulated in tumor tissues compared with normal tissues.The expression of TMEM199 is closely associated with poor prognosis of LIHC,SKCM,UCEC,and UVM,and TMEM199 may play a promoting role in tumor progression.The increased expression of TMEM199 in LUAD,LUSC,UVM,and other tumors may be related to neutrophil infiltration.The expression of TMEM199 in LAML,SARC,UCEC,and UVM were significantly positively correlated with the expression of immune checkpoint genes such as PD-L1 and LAG3.The expression of TMEM199 was positively correlated with TMB and MSI in LIHC and UCEC.Conclusion: TMEM199 induces the activation of the Canonical Wnt signaling pathway and EMT process of lung adenocarcinoma cells,and promotes their proliferation and invasion ability.TMEM199 can be used as a prognostic biomarker for pan-cancer and is associated with immune infiltration.