HSP70 binds RNA and regulates transcription by RNA Polymerase III [ChIP-seq]

We examined the RNA binding ability of the 70-kDa heat shock protein (HSP70) family member HSPA1A and identified the RNA species bound to HSP70 in vivo. We used a UV-crosslinkiung and immunoprecipitation based approach named FLASH-seq (Aktaş et al., 2017) using 3xFLAG-Histidine-Biotin tagged ectopically expressed protein in HEK293T cells. HEK293 Flp-In T-Rex with stable integration of pcDNA5-FRT-TO_3xFLAG-HBH_GFP and pcDNA5-FRT-TO_3xFLAG-HBH_HSPA1A were obtained by Flp-mediated recombination. Expression of 3xFLAG-HBH_GFP and 3xFLAG-HBH_HSPA1A proteins was induced with 1 g/ml tetracycline for 16 hours. ChIP-seq of 3xFLAG-HBH tagged GFP and HSPA1A