Functional Study Of PRDM14 On Cell Proliferation And Apoptosis Of Non-small Cell Lung Cancer Cell Line A549

The PRDM(PRDI-BF1 and RIZ homologous domain-containing proteins)family contains novel transcriptional regulators that share a characteristic molecular structure,with C-terminal zinc finger repeats and an N-terminal PR(PRDI-BF1 and RIZ homologous)domain,that shares homology with the SET(Su(var)3-9 enhancer-of-zeste and trithorax)domain.As a member of PRDM family,PRDM14 can participate in protein interaction and gene expression regulation.Abundant reports have shown that PRDM 14 plays important roles in primordial germ cell specification,embryonic stem cell pluripotency,epigenetic reprogramming.It is also known to be a key regulator of cell differentiation,growth and apoptosis.In addition,PRDM 14 has been identified as a susceptibility locus for several types of cancer.However,little is known about the mechanisms by which PRDM 14 functions in cancer development and chemoresistance of cancer cells.PRDM14 is located on 8q13,a region where gene amplification has frequently been detected in patients with non-small cell lung cancer(NSCLC)and PRDM14 has a high expression in NSCLC tissues compared with that in normal tissues.These indicate that PRDM 14 may involve in tumorigenesis of non-small cell lung cancer.In our studies,we constructed lentiviral expression vector or CRISPR-Cas9 vector to change the expression of PRDM14 in A549(NSCLC)cell line.To examine the effects of PRDM14 on cell proliferation,CCK8 assay,cell counting assay and cell colony formation assay were performed in vitro and nude mouse tumorigenicity assay was confirmed in vivo.Our results showed that PRDM 14 could promote cell proliferation.To understand the mechanisms by which PRDM 14 functions on cell proliferation,we detected the cell cycle and cell cycle regulatory factors expression levels via flow cytometric analysis and qRT-PCR.The results showed that PRDM14 potentially promoted A549 cell cycle by influencing the transcript levels of cell cycle regulatory factors.In the A549 cells overexpressing PRDM 14,the levels of the cyclin B1,cyclin DI,cyclin E1,CDK4 and CDK6 transcripts were up-regulated compared with the control.However,the level of the CKI p21 transcript was decreased compared with the control.To further investigate whether PRDM 14 can impact the A549 cells metastasis and invasion,we detected cell migration abilities via wound healing assay and transwell assay.The results showed that PRDM14 could significantly enhance metastasis and invasion abilities of A549 cell line.In addition,the MMP/TIMP(Matrix metalloproteinase/Tissue inhibitor of metalloproteinase)expression level was detected by qRT-PCR.The results showed that PRDM 14 promoted A549 cells migration mainly depending on up-regulating the expression of MMP2/TIMP2.In order to define the role of PRDM14 on drug resistance,we used Taxol,DDP,VCR and VDS to treat A549 cells and investigated apoptosis cells via flow cytometric analysis.The results showed that PRDM 14 could inhibit cell apoptosis induced by chemotherapy drugs,resulting in decreasing the sensitivity to chemotherapy for A549 cells.To further understand the mechanisms by which PRDM14 on cell proliferation and apoptosis,we detected the expression of various proteins involved in lung cancer.Results showed that PRDM14 overexpression significantly promoted the activation of p38 MAPK and NF-κB,and PRDM14 could affect the expression of CKI p27 and tumor inhibit protein p53.Taken together,our results suggested that PRDM14 could promote A549 cell proliferation,migration and invasion abilities,and it could regulate the expression levels of cell cycle regulators to affect cell cycle progression.Moreover,PRDM14 could inhibit A549 cell apoptosis induced by chemotherapy drugs via p38 MAPK signal pathway to reduce their sensitivity to chemotherapeutic drugs.Theses results indicate that PRDM14 may play an important role in NSCLC cell proliferation and it could be an ideal therapeutic target for the treatment of NSCLC.