Wilms' tumor suppressor gene (WT1) suppresses apoptosis by transcriptionally downregulating BAX expression in immature rat granulosa cellsopen access
- Authors
- Park, Minji; Choi, Yuri; Choi, Hyeonhae; Roh, Jaesook
- Issue Date
- Dec-2014
- Publisher
- BIOMED CENTRAL LTD
- Keywords
- WT1; Apoptosis; Bax; bcl-2; Caspase; Granulosa cell; Rat
- Citation
- JOURNAL OF OVARIAN RESEARCH, v.7, pp.1 - 10
- Indexed
- SCIE
SCOPUS
- Journal Title
- JOURNAL OF OVARIAN RESEARCH
- Volume
- 7
- Start Page
- 1
- End Page
- 10
- URI
- https://scholarworks.bwise.kr/hanyang/handle/2021.sw.hanyang/158502
- DOI
- 10.1186/s13048-014-0118-y
- ISSN
- 1757-2215
- Abstract
- Background: The important role of WT1 in early folliculogenesis was evident from its restricted expression pattern in immature follicles and from its involvement in transcriptional control of inhibin-a and FSH receptor. There is also considerable evidence that WT1 is a potent inhibitor of apoptotic cell death in the developing kidney and male germ cells, suggesting that it could play a role in the regulation of follicle survival. Therefore, we evaluated if WT1 involves in cell survival of granulosa cells (GCs) during the FSH-independent stage.
Methods: GCs were obtained from small preantral follicles of immature rat ovary. Bax and bcl-2 mRNA and protein levels in GCs transfected with WT1 (-KTS) or WT1 (+KTS) were analyzed by Real-time RT-PCR and immune-blotting analysis. Cell viability was measured with MTT assays and apoptosis was analyzed with caspase 3/7 activity and TUNEL assay. The mechanism by which WT1 regulates Bax expression was investigated using Bax promoter-luciferase reporter assay and ChIP assays from GCs.
Results: Here, we showed that WT1 (-KTS) suppressed endogenous Bax transcript and protein expression, and this inhibition resulted from direct binding of WT1 in the Bax promoter region in vivo. In addition, anti-apoptotic effects of WT1 (-KTS) were demonstrated based on MTT assays, a sensitive bioluminescence caspase 3/7 assay and TUNEL assays. On the other hand, WT1 has no role on bcl-2 expression in GCs.
Conclusion: These findings suggest that activation of WT1 is necessary for maintenance of GC survival during early stage of follicles and WT1 can play a role in protecting apoptosis through the regulation of upstream activator (Bax), as well as through regulation of downstream effecter (caspases 3 and 7).
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