Gene expression profiling of ISWI chromatin remodeler mutants before and after DNA damage

Gene expression profiling of ISWI chromatin remodeler mutants before and after DNA damage BAZ1A is a regulatory subunit within the ISWI family of chromatin remodelers in humans. We and others have shown that BAZ1A is required for efficient recovery from DNA damage. The DNA damage hypersensitivity caused by BAZ1A* is as severe as the deletion of the entire gene. This is striking given that BAZ1A* contains only two substitutions (K1181A and K1183Y) within a poorly characterized domain of BAZ1A. We find that this domain can interact with DNA, and that the mutations present in BAZ1A* hinder DNA binding. We used transcriptional profiling to identify molecular events leading to the DNA damage hypersensitivity of BAZ1A* cells. HeLa cells (CL132529) were subjected to genome editing using the Cas9 technology to abrogate expression of the chromatin regulator BAZ1A. Lentiviral transduction was then used to re-express either BAZ1A wild-type or the BAZ1A* mutant in BAZ1A knockout cells. All genome edited cell lines have been cloned and passed in-house QC (mycoplasma-free and cell line authenticated). Cells were either unchallenged (control) or subjected to UVC irradiation (200 J/m2) to cause DNA damage. One hour after irradiation, total RNA was extracted in triplicate. Gene expression was measured by RNA-sequencing. In total there are 8 conditions: 4 cell lines (parental (HeLa), BAZ1A knockout, re-express BAZ1A, re-express BAZ1A* mutant), with or without DNA damage. Each condition includes triplicate RNA extractions, for a total of 24 samples.