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Published on: December 26, 2016
A TALEN-based specific transcript knock-down of PIWIL2 suppresses cell growth in HepG2 tumor cell
1Department of Medical Genetics, State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, 610000, China.
Objectives:
PIWIL2 is widely expressed in various tumours and implicated in playing a role in tumourigenesis. For a more thorough study of PIWIL2 functions in tumour cells, we aimed to establish PIWIL2-specific transcript knock-down/knockout HepG2 cell lines using transcription activator-like effector nuclease (TALEN) technology. Furthermore, we proposed to use the cell models to explore PIWIL2 functions in TGF-β signalling in HepG2 cells. HepG2s are human hepatocellular carcinoma cells.
Materials And Methods:
We established PIWIL2 knock-down or knock-out cell lines in HepG2 using TALEN technology. Next, we sought to use the cell line models to investigate effects of full length PIWIL2-specific transcripts in cell proliferation induced by TGF-β.
Results:
First, we established PIWIL2-specific transcript mono-allele and bi-allele knockout HepG2 cell lines. By using the cell line models, we found that specific transcript knockdown of full length PIWIL2 can suppress cell proliferation, while ectopic expression of PIWIL2 enhanced proliferation of HepG2 by suppressing the TGF-β pathway. Furthermore, we demonstrated that PIWIL2 can interact with HSP90 to prevent formation of HSP90-TβR complexes, which promote degradation of TβRs.
Conclusions:
Taken together, our study revealed critical negative regulation of TGF-β signalling by PIWIL2 in HepG2 tumour cells, and provided an effective strategy to study specific gene transcript functions in cells.
Insights
PIWIL2 knockdown suppresses hepatocellular carcinoma cell proliferation by inhibiting TGF-β signaling. This study provides a new method for studying gene functions in cancer cells.
Area of Science:
- Cancer Biology
- Molecular Oncology
- Gene Regulation
Background:
- PIWIL2 is frequently expressed in tumors and contributes to tumorigenesis.
- Understanding PIWIL2's role in cancer requires specific cellular models.
- HepG2 cells, a human hepatocellular carcinoma line, are used to study PIWIL2.
Purpose of the Study:
- To create PIWIL2-specific knock-down/knock-out HepG2 cell lines using TALEN technology.
- To investigate the function of PIWIL2 in TGF-β signaling pathways within HepG2 cells.
- To elucidate the mechanism by which PIWIL2 influences hepatocellular carcinoma cell proliferation.
Main Methods:
- Development of PIWIL2 mono-allele and bi-allele knockout HepG2 cell lines via TALEN.
- Assessment of PIWIL2 transcript knockdown effects on HepG2 cell proliferation.
- Analysis of PIWIL2's interaction with HSP90 and its impact on TGF-β receptor (TβR) stability.
Main Results:
- Established PIWIL2 knockout HepG2 cell lines.
- PIWIL2 knockdown suppressed HepG2 cell proliferation.
- PIWIL2 overexpression enhanced proliferation by suppressing TGF-β signaling, interacting with HSP90 to prevent TβR degradation.
Conclusions:
- PIWIL2 negatively regulates TGF-β signaling in HepG2 hepatocellular carcinoma cells.
- This research offers an effective strategy for studying specific gene transcript functions.
- Targeting PIWIL2 may represent a therapeutic approach for hepatocellular carcinoma.
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