A Study On The Role Of DAP1 In Apoptosis Of Human Lung Cancer Cells

DAP1(Death associated protein 1)is an evolutionarily highly conserved protein with a molecular weight of 15 kDa,which plays an important role in cells apoptosis in the previous study,and is supposed to one of the criteria for evaluating the prognosis of cancer patients.However,it is unclear that the detailed molecular mechanism of DAP1 on apoptosis.Therefore,our aim is to explore the role of DAP1 in lung cancer cells apoptosis and the molecular mechanism.This will provide theoretical basis and new therapeutic targets for the treatment of lung cancer,and provide a good reference for the treatment of other cancers.Firstly,when we treated human lung cancer cells with chemotherapy drugs of doxorubicin and pemetrexed,we found that DAP 1 was upregulated by the drugs and is induced in a dose and time-dependent manner.Then we knocked down DAP1 and performed western blot and flow cytometry analysis,we found that the knockdown of DAP1 obviously decreased apoptosis induced by chemotherapy drugs.When we overexpressed DAP1,the apoptosis induced by chemotherapy drugs was enhanced significantly.These data reveales that DAP1 promotes lung cancer cells apoptosis.Because many chemotherapeutic drugs can induce apoptosis through endoplasmic reticulum stress,we detected the receptor protein of endoplasmic reticulum stress and found that EIF2AK3 was obviously regulated by DAP1.DAP1 suppression could dramatically down-regulate EIF2AK3-DDIT3 pathway,and DAP1 overexpression up-regulated this pathway distinctly.This indicated that DAP1 promoted ER stress by the regulation of EIF2AK3.To explore the relation of ER stress and apoptosis,we detected the ER stress associated proteins:EIF2AK3 and DDIT3,and apoptosis related proteins such as TNFRSF10B?CASP8?CASP3 and PARP-1.We found that DAP1 could promote ER stress and apoptosis at the same time.Then we overexpressed DAP1 meanwhile knocked down EIF2AK3,we found that the expression of DAP1 could not affect the apoptosis obviously.The above experimental data showed that DAP1 could induce ER stress through EIF2AK3-DDIT3 pathway,and DDIT3 induced extrinsic apoptosis by promoting the transcription of death receptor TNFRSF10B.Then We discovered the interaction between DAP1 and SETDB1 by immune coprecipitation experiment.The regulation of EIF2AK3 by DAP1 was SETDB1-dependent which was in the transcriptional leval via RT-PCR and the promoter activity analysis of EIF2AK3.In summary,We put forward the role of DAP1 in apoptosis of human lung cancer cells.Namely,DAP1 promoted the transcription of EIF2AK3 through interacting with histone methyltransferase SETDB1 which could enhance Tri-methylation of H3K4,then induced ER stress through EIF2AK3-DDIT3 pathway,and DDIT3 induced extrinsic apoptosis by promoting the transcription of death receptor TNFRSF10B.