Low-Dose-Rate Radiation-Induced Secretion of TGF-β3 Together with an Activator in Small Extracellular Vesicles Modifies Low-Dose Hyper-Radiosensitivity through ALK1 Binding

Hyper-radiosensitivity (HRS) is the increased sensitivity to low doses of ionizing radiation observed in most cell lines. We previously demonstrated that HRS is permanently abolished in cells irradiated at a low dose rate (LDR), in a mechanism dependent on transforming growth factor beta 3 (TGF-beta 3). In this study, we aimed to elucidate the activation and receptor binding of TGF-beta 3 in this mechanism. T-47D cells were pretreated with inhibitors of potential receptors and activators of TGF-beta 3, along with addition of small extracellular vesicles (sEVs) from LDR primed cells, before their radiosensitivity was assessed by the clonogenic assay. The protein content of sEVs from LDR primed cells was analyzed with mass spectrometry. Our results show that sEVs contain TGF-beta 3 regardless of priming status, but only sEVs from LDR primed cells remove HRS in reporter cells. Inhibition of the matrix metalloproteinase (MMP) family prevents removal of HRS, suggesting an MMP-dependent activation of TGF-beta 3 in the LDR primed cells. We demonstrate a functional interaction between TGF-beta 3 and activin receptor like kinase 1 (ALK1) by showing that TGF-beta 3 removes HRS through ALK1 binding, independent of ALK5 and TGF-beta RII. These results are an important contribution to a more comprehensive understanding of the mechanism behind TGF-beta 3 mediated removal of HRS.

Automated summary

This study aimed to elucidate the mechanism of transforming growth factor beta 3 (TGF-beta 3) activation and receptor binding in low dose rate (LDR) irradiated cells. The results showed that small extracellular vesicles (sEVs) from LDR primed cells could remove hyper-radiosensitivity (HRS), and inhibition of the matrix metalloproteinase (MMP) family prevented the removal of HRS. Additionally, functional interaction between TGF-beta 3 and activin receptor like kinase 1 (ALK1) was demonstrated in the removal of HRS, independent of ALK5 and TGF-beta RII.