Medizinische Universität Graz Austria/Österreich - Forschungsportal - Medical University of Graz
Gewählte Publikation:
Pocha, J.
Modulation of AKT signalling by the gluconeogenesis enzyme PCK2 in lung cancer cells
Humanmedizin; [ Diplomarbeit ] Medical University of Graz; 2021. pp. 52
[OPEN ACCESS]
FullText
- Autor*innen der Med Uni Graz:
- Betreuer*innen:
- Leithner Katharina
- Altmetrics:
- Abstract:
- Despite extensive preventive measures, lung cancer remains the cancer with the highest mortality, demanding further research for therapeutic targets. One potential therapeutic approach is the remodelling of energy metabolism. Changing supplies of oxygen and nutrients require high metabolic flexibility. Leithner et al. in 2014 showed activity of the gluconeogenesis pacemaker enzyme phosphoenolpyruvate carboxykinase (PCK2) in non-small-cell lung carcinoma (NSCLC) cells, which thereby increases the metabolic flexibility. However, it is unknown whether this modulates signal transduction in tumour cells. This study aimed to evaluate the effects of PCK2 gene silencing via siRNA on signalling pathways affecting cell growth. The PI3K/AKT/mTORC signalling pathway was especially of interest due to the role of mTORC1 and mTORC2 as central signalling hubs and the mediation of signalling related to survival, proliferation, autophagy, cell cycle and growth. Cells of the two NSCLC cell lines A549 and H23 were cultured and transfected with siRNA under high and low glucose conditions. Phosphorylation levels of AKT (S473), AKT (T308), AMPK and S6K were analysed in Western blots. Furthermore, the expression of various membrane transporters and the IGF-1 receptor was evaluated with qPCR. Silencing of PCK2 led to increased phosphorylation of AKT (S473) in H23 under high (p = 0.012; <0.05) as well as under low glucose conditions in the medium. The use of different inhibitors allowed to attribute this to increased mTORC2 activation under PCK2 inhibition. No distinct effects on AKT T308, S6K or AMPK could be observed by silencing PCK2, albeit cell line dependent effects of glucose concentration were observed. PCK2 silencing in H23 cells showed overexpression in the qPCR of GLUT1 (SLC2A1) and the glutamine transporter (SLC1A5) under high glucose medium conditions, possibly as a means of adaptation. Altogether, this study's results suggest modulation of the mTORC2 pathway through the activity of the gluconeogenesis enzyme PCK2, although highly dependent on individual cell lines and respective properties. In the H23 cell line, mutant TP53 could play a role in effects on AKT (S473) observed in the experiments.