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P1.03 - Biology (Not CME Accredited Session) (ID 935)
- Event: WCLC 2018
- Type: Poster Viewing in the Exhibit Hall
- Presentations: 1
- Coordinates: 9/24/2018, 16:45 - 18:00, Exhibit Hall
P1.03-17 - Functional Mechanism of GLDC Gene Alternative Splicing in Non-Small Cell Lung Cancer (ID 12270)
16:45 - 18:00 | Author(s): Yingli Yuan
Lung cancer is the leading cause of death in the world and non-small cell lung cancer (NSCLC) accounts for approximately 85%.While, the mechanism is not fully understood.Therefore,exploring new molecules is of great significance to clarify the mechanism.Glycine decarboxylase (GLDC) is an oncogene associated with lactic acid metabolism,highly expressed in cancer stem cell-riched NSCLC.Both growth and tumorigenicity are dependent on the expression of high levels of GLDC, so which can be used as a stem cell surface marker to identify cancer stem cells.Alternative splicing (AS) is one of the important mechanisms of human gene regulation,which is closely related to the occurrence of tumor.However,the mechanism of AS regulation for GLDC in NSCLC has not been reported.Therefore,understanding the biological role of GLDC and its regulatory mechansim can lay a theoretical foundation for the early diagnosis and treatment of tumors.This experiment aims to clone human GLDC gene AS,analyze biological activity,and provide guidance for NSCLC development mechanisma9ded1e5ce5d75814730bb4caaf49419 Method
1.Construction of cloning vectors:Primers were designed based on the full length of GLDC (GLDCfl) and GLDC varient 1 (GLDCV1) cDNA,and mRNA expression in A549 and MRC5 cells was determined by RT-PCR combined with DNA sequencing;4c3880bb027f159e801041b1021e88e8 Result
2.Functional experiments:MRC5 cells were transfected with overexpression plasmids,MTT assay,plate clone,western blot,lactic acid formation assay,and tumor-bearing test in vivo were used to detect MRC5 cells viability and proliferation.
1.Both GLDCfl and GLDCV1 of A549 and MRC5 existed at the RNA level,the expression of which only in A549.Transfection of the target gene in MRC5 cells showed increased expression of both GLDCfl and GLDCV1.
2.GLDCfl and GLDCV1 were over-expressed in MRC5 cells,and the cell viability was increased by MTT assay.Plate clone experiments proved the proliferation of MRC5 cells,and the expression of P-ERK and P-P38 proteins on the ERK/MAPK signaling pathway were increased.The supernatants of the cells for 24 hours were collected and assayed for lactic acid,then both GLDCV1 and GLDCfl promoted lactic acid production. Meanwhile,in vivo nude mice tumor test,each 100 μl dose was inoculated subcutaneously at the back shoulder of 4-6 week old nude mouse,and the GLDCV1 group shown the tumorigenicity8eea62084ca7e541d918e823422bd82e Conclusion
1This study first analyzed the expression pattern of GLDC gene AS in tumor cells and revealed the biological effects in tumor cells;
2.Both GLDCV1 and GLDCfl can enhance the cell viability,promote luciferase activity,enhance cell clonal formation,and also find that GLDCV1 can enhance the tumorigenic ability.
3.Both GLDCV1 and GLDCfl have effect on the expression of ERK/MAPK pathway protein.6f8b794f3246b0c1e1780bb4d4d5dc53