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Published on: September 16, 2019
WNK2 Inhibits Autophagic Flux in Human Glioblastoma Cell Line
Ana Laura Vieira Alves1, Angela Margarida Costa2,3, Olga Martinho1,2,3
1Molecular Oncology Research Center, Barretos Cancer Hospital, 14784 400 Barretos, Brazil.
Abstract:
Autophagy is a cell-survival pathway with dual role in tumorigenesis, promoting either tumor survival or tumor death. WNK2 gene, a member of the WNK (with no lysine (K)) subfamily, acts as a tumor suppressor gene in gliomas, regulating cell migration and invasion; however, its role in autophagy process is poorly explored. The WNK2-methylated human glioblastoma cell line A172 WT (wild type) was compared to transfected clones A172 EV (empty vector), and A172 WNK2 (WNK2 overexpression) for the evaluation of autophagy using an inhibitor (bafilomycin A1-baf A1) and an inducer (everolimus) of autophagic flux. Western blot and immunofluorescence approaches were used to monitor autophagic markers, LC3A/B and SQSTM1/p62. A172 WNK2 cells presented a significant decrease in LC3B and p62 protein levels, and in LC3A/B ratio when compared with control cells, after treatment with baf A1 + everolimus, suggesting that WNK2 overexpression inhibits the autophagic flux in gliomas. The mTOR pathway was also evaluated under the same conditions, and the observed results suggest that the inhibition of autophagy mediated by WNK2 occurs through a mTOR-independent pathway. In conclusion, the evaluation of the autophagic process demonstrated that WNK2 inhibits the autophagic flux in glioblastoma cell line.
Insights
WNK2 gene overexpression inhibits autophagy in glioblastoma cells. This finding suggests WNK2 may be a therapeutic target for glioblastoma by modulating the autophagic process.
Area of Science:
- Cell Biology
- Oncology
- Molecular Biology
Background:
- Autophagy is a critical cellular process involved in cancer development, with roles in both tumor survival and death.
- The WNK2 gene functions as a tumor suppressor in gliomas, affecting cell migration and invasion.
- The specific role of WNK2 in the autophagy pathway within gliomas remains largely uninvestigated.
Purpose of the Study:
- To investigate the role of WNK2 in regulating autophagy in human glioblastoma cells.
- To determine if WNK2 overexpression impacts autophagic flux.
- To explore the potential involvement of the mTOR pathway in WNK2-mediated autophagy regulation.
Main Methods:
- Utilized human glioblastoma cell lines: A172 WT, A172 EV (empty vector control), and A172 WNK2 (WNK2 overexpression).
- Assessed autophagic flux using bafilomycin A1 (autophagy inhibitor) and everolimus (autophagy inducer).
- Monitored autophagy markers LC3A/B and SQSTM1/p62 via Western blot and immunofluorescence.
Main Results:
- WNK2 overexpression in A172 cells led to a significant reduction in LC3B and p62 protein levels.
- The ratio of LC3A/B was decreased in WNK2-overexpressing cells compared to controls after treatment.
- These results indicate that WNK2 overexpression inhibits autophagic flux in glioblastoma cells.
- WNK2-mediated inhibition of autophagy appears to operate independently of the mTOR pathway.
Conclusions:
- WNK2 functions to inhibit autophagic flux in glioblastoma cells.
- The findings suggest a novel mechanism by which WNK2 influences glioblastoma biology.
- WNK2's role in modulating autophagy presents a potential therapeutic avenue for glioblastoma treatment.
