A heterogeneous cellular response to ionizing radiation revealed by single cell transcriptome sequencing.

Our understanding on transcriptional regulation of tumour cells responding to ionizing radiation (IR) has mostly come from bulk sequencing. However, due to the heterogeneity of tumour, how each individual cell responds to IR differently is unclear. We report here a heterogeneous cellular response to IR by single cell transcriptome sequencing. We utilized the barcoded Smart-seq2 single cell transcriptome sequencing technology in breast cancer cell line MDA-MB-231 both without and with IR treatment. To further understand how ATM, a major hub protein required for an optimal DNA damage response, affected the heterogeneous IR response, we also knocked down ATM gene for single cell transcriptome sequencing. Single cell t-SNE analysis showed four clusters of cells responding to IR in distinctive ways: Cluster 1 changed the least; Cluster 2 responded to IR by upregulating ribosome associated genes, while Cluster 4 upregulated both ribosome and G1/S phase associated genes; Cluster 3 was a new cluster, which appeared only in irradiated cells. In the absence of ATM kinase, cells displayed much less transcriptional changes after IR. And Cluster 4 in wild-type cells, which had the greatest change in response to IR, was not present in the ATM knock-down cells. We also selected three IR-induced genes for functional validation in both MDA-MB-231 and an additional breast cancer cell line to demonstrate their importance in radiation sensitivity. Taken together, our single cell transcriptome analysis has revealed a heterogeneous cellular response to DNA damage induced by IR and identified potential biomarkers of radiation sensitivity.