Exosome-packaged miR-1246 contributes to bystander DNA damage by targeting LIG4

Abstract

Background An increasing number of studies have recently reported that microRNAs packaged in exosomes contribute to multiple biological processes such as cancer progression; however, little is known about their role in the development of radiation-induced bystander effects. Methods The exosomes were isolated from the culture medium of BEP2D cells with or without γ-ray irradiation by ultracentrifugation. To monitor DNA damage and repair efficiency, the DNA double-strand break biomarker 53BP1 foci, comet, micronuclei, expression of DNA repair genes and NHEJ repair activity were detected. The miR-1246 targeting sequence of the DNA ligase 4 ( LIG4 ) mRNA 3′UTR was assessed by luciferase reporter vectors. Results miR-1246 was increased in exosomes secreted from 2 Gy-irradiated BEP2D cells and inhibited the proliferation of nonirradiated cells. The miR-1246 mimic, exosomes from irradiated cells, and radiation-conditioned cell culture medium increased the yields of 53BP1 foci, comet tail and micronuclei in nonirradiated cells, and decreased NHEJ efficiency. miR-1246 downregulated LIG4 expression by directly targeting its 3′UTR. Conclusions Our findings demonstrate that miR-1246 packaged in exosomes could act as a transfer messenger and contribute to DNA damage by directly repressing the LIG4 gene. Exosomal miR-1246 may be a critical predictor of and player in radiation-induced bystander DNA damage.

Integrated analysis of transcriptomic and metabolomic profiling reveal the p53 associated pathways underlying the response to ionizing radiation in HBE cells

“…qRT-PCR was carried out in our laboratory using the Super Real PreMix Plus (SYBR Green) kit (Tiangen Biotech, Beijing, China) on the CFX96TM Real-Time system (Bio-Rad Laboratories, Hercules, CA, USA), to validate the changes and trends observed in the microarray analysis. The comparative Ct (2 −ΔΔCt ) method was used to calculate relative fold differences in mRNA expression between HBE p53-/-and HBE p53-wt cells [32,33]. mRNA expression levels were compared using the two-tailed Student's t-test.…”

“…To ascertain whether p53 deficiency affected mRNA expression post-radiation, we compared mRNA expression levels between the p53-/-HBE and p53-wt HBE cells. We conducted this analysis 24 h after 4 Gy γ radiation exposure, because human lung branchial epithelial cells exhibit 4 Gy-induced radiation injury (i.e., significant DNA damage) at this timepoint, based on our previous studies [31,33,[41][42][43]. According to the transcriptomic data, 326 mRNAs showed significantly altered expression in HBE p53-/-cells postradiation, of which 269 were upregulated and 57 were downregulated ( Fig.…”

Transcriptomic and Metabolomic Profiling Reveal the p53-Dependent Benzeneacetic Acid Attenuation of Silica-Induced Epithelial–Mesenchymal Transition in Human Bronchial Epithelial Cells

“…WB detection was conducted as previously reported [22,28,29]. In this study, antibodies against p53, E-cadherin, N-cadherin, and vimentin were purchased from Cell Signaling Technology, Inc.(Danvers, MA, USA) and Abcam (Cambridge, UK).Images were captured and assessed using the ChemiDocXRS + system (Bio-Rad Laboratories).…”

Regulation of DNA Double-Strand Break Repair by Non-Coding RNAs

“…Exosomes have been reported to contain as many as 764 miRNAs [174,175]. The miR-1246 is an example of a radiation-induced miRNA that is packaged into exosomes and is delivered to non-irradiated cells to decrease NHEJ efficiency by targeting the 3' UTR of LIG4 [155]. These diverse roles of miRNAs in the DNA damage response and cancer have made them attractive targets for cancer therapy.…”