The Effect Of PDLIM5 And BCAT1 In Non-small Cell Lung Cancer And The Potential Molecular Mechanism

Objective:Incidence and mortality of lung cancer have remained high in recent years.There were approximately 2.1 million new cases of lung cancer and 1.76 million deaths worldwide in 2018.Deaths caused by lung cancer accounted for nearly one-fifth?18.4%?of all deaths caused by cancers^[1].Non-small cell lung cancer is the most common histological type of lung cancer.In recent years,the survival time of patients with advanced NSCLC have been greatly improved in spite of the improvement of gefitinib,erlotinib and other targeted drugs into clinical application.The pathogenesis of lung cancer still needs further studies and it is essential to find early diagnosis methods and targeted drugs for lung cancer.PDLIM5 is a member of the PDZ-LIM protein family.PDZ and LIM domains mediate the interaction between proteins.The amino terminus of PDLIM5 contains a PDZ domain,and the carboxyl terminus of PDLIM5 contains three LIM domains.Previous studies have confirmed that PDLIM5 regulats the signal transduction of membrane related proteins and cytoskeletal proteins^[2-4].The abnormal expression of PDLIM5 was observed in breast cancer,gastric cancer and prostate cancer^[5-8].PDLIM5can regulate tumor progression.However,the mechanism of PDLIM5 in NSCLC has not been reported.Branched-chain amino acid transaminases?BCATs?can catalyze the transamination of branched-chain amino acids and convert them into branched-chain?-pyruvates and glutamates^[9].BCATs contains two important isoenzymes,cytoplasmic branched chain amino acid transferase?BCTA1?and mitochondrial branched chain amino acid transferase?BCAT2?^[10].They both played important roles in many tumors.It has been reported that BCAT1 was up-regulated in liver cancer,breast cancer and cervical cancer.BCAT1 promoted proliferation and invasion in these tumors^[11-13].However,the mechanism of BCAT1 in lung cancer remains unclear.In this study,we analyzed the expression of PDLIM5 and BCAT1 in lung cancer tissues and cell lines.The clinical significance of PDLIM5 and BCAT1 was also analyzed.The biological function and potential molecular mechanism of PDLIM5 and BCAT1 in lung cancer cells were further investigated.Methods:The expression of PDLIM5 and BCAT1 in human lung cancer tissues and the corresponding normal tissues was detected by immunohistochemistry.The clinical significances was also analyzed.Effects of PDLIM5 and BCAT1 on proliferation in NSCLC cells were explored by MTT assay and colony formation assay.Transwell matrigel invasion assay was used for determining the effect of PDLIM5 and BCAT1 on the invasion in non-small cell lung cancer cells.The influence of PDLIM5 on apoptosis induced by erlotinib in non-small cell lung cancer cells was determined by flow cytometry.RT-PCR and western bolt were used to detect expressions of BCAT1 and PDLIM5 in normal alveolar epithelial cells and non-small cell lung cancer cells.The influence of PDLIM5 on proteins related to proliferation and Hippo signaling pathway was explored.The influences of BCAT1 on Wnt/?-catenin signaling pathway was also detected using western blot.SPSS 16.0 software was used for statistical analysis.The overall survival was evaluated by Kaplan-Meier method,and the survival difference was verified by log-rank test.Data of experimental group and control group were analyzed by Student's t-test.P<0.05 indicates statistical significance.Results:1.Immunohistochemistry results showed that the expression of PDLIM5increased significantly in lung cancer tissues compared with normal tissues.PDLIM5high level was significantly correlated with advanced TNM stage?p=0.002?,tumor status?p=0.0013?and lymph node metastasis?p=0.001?in patients with non-small lung cancer.Western blot results showed that the expression of PDLIM5 in non-small-cell lung cancer cells was significantly higher than normal cells.MTT assay and colony formation assay results showed that PDLIM5 promoted proliferation and colony formation in non-small cell lung cancer cells.Flow cytometry results showed that PDLIM5 inhibited erlotinib-induced apoptosis in non-small cell lung cancer cells.The results of transwell matrigel invasion assay showed that PDLIM5 significantly improved the invasion ability in non-small cell lung cancer cells.Western blot results showed that PDLIM5overexpression up-regulated the expression of cyclin E,YAP,TAZ,and down-regulated the expression of p21 and p-LATS1.2.Immunohistochemistry results showed that BCAT1 expression increased significantly in lung cancer tissues compared with normal tissues.BCAT1 high expression was significantly correlated with advanced TNM stage?p=0.0149?and lymph node metastasis?p=0.0172?in patients.Western blot and RT-PCR results showed that the expression of BCAT1 in non-small cell lung cancer cells was significantly higher than that in normal alveolar epithelial cells.MTT assay and colony formation assay results showed that BCAT1 significantly improved the proliferation and colony formation ability of NSCLC.The results of transwell matrigel invasion assay showed that BCAT1significantly improved the invasion ability of non-small cell lung cancer cells.For the molecular mechanism,western blot results showed that BCAT1 overexpression up-regulated the expression of cylin D1,c-myc,MMP7,active-?-catenin,while down-regulated the expression of p27,E-cadherin and p-?-catenin.BCAT1 deficiency showed the opposite results.Conclusion:PDLIM5 promoted proliferation and invasion,while inhibited apoptosis induced by erlotinib in non-small cell lung cancer cells.PDLIM5 inhibited the Hippo pathway and regulateed the expressions of cyclin E,p21,YAP,TAZ and LATS1.BCAT1 promoted proliferation and invasion in non-small cell lung cancer cells.BCAT1activated the Wnt/?-catenin signaling pathway and regulated expression of cyclin D1,c-myc,MMP7 and?-catenin.