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"CONNEXINS AS POTENTIAL TARGETS FOR MODULATING LOW-DOSE
RADIATION RESPONSES IN TUMOUR VERSUS NORMAL CELLS"
Chandna Sudhir* and Ghosh Soma
Natural Radiation Response Mechanisms Group, Division of Radiation Biosciences, Institute of
Nuclear Medicine & Allied Sciences, Delhi- 110054, India; *[email protected]
Low sub-lethal doses of radiation to tumour regions cause secondary malignancies, the
mechanisms of which are not yet established. Studies also indicated connexin upregulation by
low doses while connexins are independently shown to alter cell migration in the unirradiated
cells. We hence investigated low-dose gamma-radiation induced alterations in connexin-43
(Cx43) expression as well as in cell proliferation/ migration/ invasion, along with the putative
molecular pathways such as p38 and ERK-1/2 MAPKinases. Interestingly, a narrow range of low
doses (10cGy-20cGy) enhanced Cx43 expression and also selectively induced tumour cell
migration without altering cell proliferation. Low-dose induced cell migration and p38 activation
was strongly inhibited by knocking down Cx43 expression. Silencing Cx43 caused nearcomplete inhibition of radiation-induced cell migration/invasion in all tumour cell lines (U87,
BMG-1, A549 and HeLa), whereas no cell migration/invasiveness could be induced in the
irradiated primary VH10 or transformed AA8 fibroblasts, thereby demonstrating the regulatory
role of connexins.
We further evaluated the fate of tumour and normal cells in the event of silencing connexin-43
prior to irradiation at low doses. Asynchronously growing or G2/M enriched cultures of glioma
cells (U87, BMG-1), cervical cancer cells (HeLa) and normal HDFn fibroblasts were γ-irradiated
at varying doses, with a parallel asynchronous group subjected to Cx43-knockdown prior to
irradiation. About ~60% depletion of Cx43 by siRNA elicited quite prominent HRS response,
directly implicating Cx43 in this phenomenon. In the Cx43-depleted cells, low-dose irradiation
also caused significant cell growth inhibition and apoptotic events including loss of
mitochondrial membrane potential (MMP), cytochrome-c release and caspase-3 activation
associated with Bax translocation. Therefore, downregulation of Cx43 renders tumour cells
highly susceptible to low-dose HRS. Incidentally, G2/M-phase cells have significantly reduced
levels of adhesion proteins such as Cx43 and also display enhanced low-dose hyperradiosensitivity (HRS). G2/M enrichment indeed caused considerable HRS at doses 10cGy30cGy in all tumour cell lines and was associated with significantly reduced (~50%) Cx43 level.
Our study thus demonstrates that Cx43 mediates low-dose radiation induced cell migration
selectively in tumour cells, and silencing it reverses this process and renders tumour cells
hypersensitive to low doses. The study may have strong implications for therapeutics.