Transcriptional readthrough and splicing upon chemotherapy treatment

The Neugebauer lab has previously shown that splicing occurs co-transcriptionally and is coupled to 3'end cleavage. It is known that stress, such as hyperosmotic shock, affects RNA processing. The cellular stress response is an important component of cancer cell response to chemotherapy that can determine cell survival. Herein, we study how the coupling of the RNA processes in leukemia cells is affected in response to stress and therapeutic agents. We used short- and long-read sequencing to follow RNA processing upon treatment. Overall design: To determine level of gene expression and transcriptional readthrough as well as determine changes in alternative splicing we used short-read RNA-seq. We analyzed polyA selected RNA isolated from whole K562 cells (mRNA) and RNA isolated from the cell nucleuses (nuclear RNA) that was later rRNA-depleted. Before isolation of total RNA the cells were treated with: - 1uM Imatinib (im) for 1h, 6h and 18h - 5uM JTE-607 (jte) for 1h, 6h and 18h - 110mM KCl (kcl) for 1h and 3h. Before isolation of nuclear RNA cells were treated with: - im for 1h or 18h - jte for 1h - kcl for 1h All time points ended at the same time and thus the cells were collected in paralell with the untreated control cells (ctrl). Each experiment was repeated three times (1, 2, 3)