- Authors: Gilardini Montani MS. et al.
- Year: 2021
- Journal: Carcinogenesis
- Applications: in vitro / siRNA / INTERFERin
- Cell type: B cells
Briefly, the day of transfection, lymphocytes were seeded into 6-well plates at a density of a 3x10^6 cells/well in 1 mL of RPMI medium without antibiotics. Next, 6 µL of 100 µM siRNA combined with 12 µL of INTERFERin were added to the cells. Transfection reactions were performed in serum-free OptiMEM medium. After 4 hours RPMI was added to each well to a final volume of 2 mL with 10% FBS. After 24 hours cells were EBV or Mock infected and cultured for further analysis.
Reactive oxygen species (ROS) and DNA repair respectively promote and limit oncogenic transformation of B cells driven by Epstein-Barr virus (EBV). We have previously shown that EBV infection reduced autophagy in primary B lymphocytes and enhanced ROS and interleukin 6 (IL-6) release, promoting B cell proliferation and immortalization. In this study, we explored the role of p62/SQSTM1, accumulated as a consequence of autophagy reduction in EBV-infected B lymphocytes, and found that it exerted a growth suppressive effect in these cells. At molecular level, we found that p62 counteracted IL-6 production and ROS increase by interacting with NRF2 and promoting mitophagy. Moreover, p62/NRF2 axis sustained the expression level of H2AX and ataxia-telangiectasia mutated (ATM), whose activation has been shown to have growth-suppressive effects during the first steps of EBV-infection, before latency is established. In conclusion, this study shows for the first time that the accumulation of p62 and the activation of p62/axis counteracted EBV-driven proliferation of primary B lymphocytes.