Abstract
Recently, we have demonstrated that microRNA-31 (miR-31) overexpression is inherent to radiation-induced cell death in the highly radioresistant Sf9 insect cells, and regulates pro-apoptotic Bax translocation to mitochondria. In the present study, we report that at sub-lethal radiation doses for Sf9 cells, miR-31 is significantly downregulated and is tightly regulated by an unusual mechanism involving p53. While ectopic overexpression of a well-conserved Sfp53 caused typical apoptosis, radiation-induced p53 accumulation observed selectively at sub-lethal doses failed to induce cell death. Further investigation of this paradoxical response revealed an intriguing phenomenon that sub-lethal radiation doses result in accumulation of a ‘hyper-phosphorylated’ Sfp53, which in turn binds to miR-31 genomic location and suppresses its expression to prevent cell death. Interestingly, priming cells with sub-lethal doses even prevented the apoptosis induced by lethal radiation or ectopic Sfp53 overexpression. On the other hand, silencing p53 increased radiation-induced cell death by inhibiting miR-31 downregulation. This study thus shows the existence of a unique radiation-responsive ‘p53 gateway’ preventing miR-31-mediated apoptosis in Sf9 cells. Since Sfp53 has a good functional homology with human p53, this study may have significant implications for effectively modulating the mammalian cell radioresistance.
Highlights
Ionizing radiation (IR) leads to double-strand DNA breaks or DSBs which activate cell-cycle checkpoints to initiate a cohort of signals leading to determination of cell fate such as cell death, damage free cell survival or even cellular transformation
Irradiating radioresistant Sf9 cells with varying doses ranging up to 3 kGy caused significant accumulation and nuclear translocation of Sfp[53], but this phenomenon was most prominent at relatively lower doses (
While p53 showed maximal accumulation following irradiation at 200 Gy (Fig. 1a), miR-31 expression was previously reported to be unusually suppressed at this dose that failed to induce caspase-3 activity or cell death[19]
Summary
Ionizing radiation (IR) leads to double-strand DNA breaks or DSBs which activate cell-cycle checkpoints to initiate a cohort of signals leading to determination of cell fate such as cell death, damage free cell survival or even cellular transformation. Tumor suppressor p53 is one of the most extensively studied DNA damage responsive proteins, which regulates cellular radiation response and is known to be frequently mutated in human tumors. Functional analysis strongly indicated that tumor-suppressive miR-31 inhibits proliferation and promotes apoptosis in the ovarian tumor cells. These effects could be observed only in cell lines with a dysfunctional p53 signaling pathway[23]. Some studies suggested the response of Sfp[53] following DNA damaging baculovirus infection[27,28], the basis of this altered ‘radiation response’ of Sfp[53] and its role in radiation-induced cell death has not been established clearly. The study shows a rather interesting role of p53 phosphorylation status in the radioresistance of Sf9 cells, which seems to work through highly selective suppression of miR-31 expression at sub lethal doses
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