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Published on: October 4, 2019
TTBK2 circular RNA promotes glioma malignancy by regulating miR-217/HNF1β/Derlin-1 pathway
Jian Zheng1,2, Xiaobai Liu1,2, Yixue Xue3,4
1Department of Neurosurgery, Shengjing Hospital of China Medical University, Shenyang, 110004, People's Republic of China.
Background:
Circular RNAs are a subgroup of non-coding RNAs and generated by a mammalian genome. Herein, the expression and function of circular RNA circ-TTBK2 were investigated in human glioma cells.
Methods:
Fluorescence in situ hybridization and quantitative real-time PCR were conducted to profile the cell distribution and expression of circ-TTBK2 and microRNA-217 (miR-217) in glioma tissues and cells. Immunohistochemical and western blot were used to determine the expression of HNF1β and Derlin-1 in glioma tissues and cells. Stable knockdown of circ-TTBK2 or overexpression of miR-217 glioma cell lines (U87 and U251) were established to explore the function of circ-TTBK2 and miR-217 in glioma cells. Further, luciferase reports and RNA immunoprecipitation were used to investigate the correlation between circ-TTBK2 and miR-217. Cell Counting Kit-8, transwell assays, and flow cytometry were used to investigate circ-TTBK2 and miR-217 function including cell proliferation, migration and invasion, and apoptosis, respectively. ChIP assays were used to ascertain the correlations between HNF1β and Derlin-1.
Results:
We found that circ-TTBK2 was upregulated in glioma tissues and cell lines, while linear TTBK2 was not dysregulated in glioma tissues and cells. Enhanced expression of circ-TTBK2 promoted cell proliferation, migration, and invasion, while inhibited apoptosis. MiR-217 was downregulated in glioma tissues and cell lines. We also found that circ-TTBK2, but not linear TTBK2, acted as miR-217 sponge in a sequence-specific manner. In addition, upregulated circ-TTBK2 decreased miR-217 expression and there was a reciprocal negative feedback between them in an Argonaute2-dependent manner. Moreover, reintroduction of miR-217 significantly reversed circ-TTBK2-mediated promotion of glioma progression. HNF1β was a direct target of miR-217, and played oncogenic role in glioma cells. Remarkably, circ-TTBK2 knockdown combined with miR-217 overexpression led to tumor regression in vivo.
Conclusions:
These results demonstrated a novel role circ-TTBK2 in the glioma progression.
Insights
Circular RNA circ-TTBK2 promotes glioma progression by sponging miR-217, leading to increased proliferation and invasion. Targeting circ-TTBK2 and restoring miR-217 shows therapeutic potential for glioma.
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- Circular RNAs (circRNAs) are a class of non-coding RNAs derived from the mammalian genome.
- The expression and function of circ-TTBK2 in human glioma remain largely unexplored.
Purpose of the Study:
- To investigate the expression profile of circ-TTBK2 in human glioma.
- To elucidate the functional role of circ-TTBK2 in glioma cell proliferation, migration, and invasion.
- To explore the molecular mechanism underlying circ-TTBK2's function, including its interaction with microRNA-217 (miR-217).
Main Methods:
- Quantitative real-time PCR and fluorescence in situ hybridization were used to assess circ-TTBK2 and miR-217 expression in glioma tissues and cells.
- Western blot and immunohistochemistry were employed to determine HNF1β and Derlin-1 expression.
- Functional assays including Cell Counting Kit-8, transwell, and flow cytometry were performed.
- Luciferase reporter assays and RNA immunoprecipitation were utilized to confirm the interaction between circ-TTBK2 and miR-217.
Main Results:
- Circ-TTBK2 was significantly upregulated in glioma tissues and cell lines, correlating with enhanced cell proliferation, migration, and invasion.
- Circ-TTBK2 functioned as a molecular sponge for miR-217, leading to decreased miR-217 levels and reciprocal negative feedback.
- MiR-217 directly targeted HNF1β, an oncogenic factor in glioma.
- Combined knockdown of circ-TTBK2 and overexpression of miR-217 demonstrated significant tumor regression in vivo.
Conclusions:
- Circ-TTBK2 plays a crucial oncogenic role in promoting glioma progression.
- The circ-TTBK2/miR-217/HNF1β axis represents a potential therapeutic target for glioma treatment.
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