Epithelial-to-mesenchymal transition (EMT) has been linked to the regulation of non-small cell lung cancer (NSCLC) progression, migration, invasion, metastasis, and chemoresistance as well. Recent studies showed that autophagy was a novel target to mediate EMT of cancer cells through regulating the migration and invasion in cancer. The DNA Damage-Regulated Autophagy Modulator 1 (DRAM1) is an evolutionarily conserved transmembrane protein and localizes predominantly to lysosomes, while also colocalizing with the autophagosome marker LC3. Previous investigation discovered that DRAM1 played an important role in regulating autophagy, inducing apoptosis of NSCLC cells. However, the function and mechanism of DRAM1 mediated autophagy in regulating EMT in NSCLC remains poorly understood. In this study, transforming growth factor-β (TGF-β) and starvation were employed to induce autophagy for investigating EMT in NSCLC A549 cells. The results showed that after exposure to TGF-β 24h, the protein levels of DRAM1, autophagy marker SQSTM1/p62, LC3I/II and EMT marker vimentin significantly increased while another EMT marker E-cadherin protein decreased obviously compared with the control by western blot. Also the similar results were observed after starvation 48h in this study. To further illustrate the role of DRAM1 in regulation of autophagy and EMT, wildtype DRAM1 overexpression plasmid and DRAM1 siRNA was transfected into A549 cells respectively to check the autophagy markers and EMT markers expression, meanwhile wound healing test and cell invasion transwell chamber test were assayed after the gene manipulation. These experiments results demonstrated that DRAM1 overexpression dramatically increased the SQSTM1/p62, LC3I/II and vimentin protein levels and inhibited the expression of E-cadherin, promoting the migration and invasion abilities of A549 cells. Conversely, down-regulation of DRAM1 restrained SQSTM1/p62, LC3 I/II and vimentin protein levels and elevated the expression of E-cadherin, leading to the inhibition of migration and invasion abilities of A549 cells. Interestingly, our further study showed that after p62 siRNA transfected into A549 cells E-cadherin upregulated while vimentin reduced compared to the control group. Consistently, knockdown of p62 suppressed the migration and invasion abilities of A549 cells and down regulated the expression of twist, which is an EMT related transcription factor. These data suggested that DRAM1 played an important role in regulating the EMT of lung cancer A549 cells through SQSTM1/p62 modulating, which might be a novel potential target for lung cancer treatment.
Citation Format: Binfeng He, Jiajing Cai, Guangcheng Huang, Fan Chang, Lei Xu, Dongsheng Wang, Yibin Deng, Xiaolan Guo. DRAM1 regulates EMT of lung cancer A549 cells via SQSTM1/p62 in vitro. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1678.