NOTCH-mediated non-cell autonomous regulation of chromatin structure during senescence [ATAC-Seq]

We describe the accessible chormatin landscape in RAS-induced (RIS) and NOTCH induced senescence (NIS) using ATAC-seq. By expressing active NOTCH (N1ICD) in the context of RIS, we find that N1ICD antagonizes the formation of accessible regions in RIS. By performing co-cultures, we demonstrate that cells expressing a NOTCH1 ligand, JAGGED1, can antagonize the formation of RIS specific accessible regions. Interrogation of accessible regions in RIS and NIS cells using ATAC-seq in IMR90 cells expressing ER:HRAS(G12V) and infected with a control (pWZL) vector or pWZL FLAG-N1ICD, and treated with 100nM 4OHT for 6 days or untreated. In co-culture experiments, IMR90 cells expressing ER:HRAS(G12V) and mRFP1 are cultured at a ratio of 1:1 with tumour cell lines for 6 days with 4OHT before isolating red (IMR90 cells) using flow cytometry to perform ATAC-seq. To determine the effect of HMGA1 depletion on the distribution of nucleosome free regions in RIS cells, IMR90 cells expressing ER:HRAS(G12V) were retrovirally infected with a short hairpin targetting HMGA1 (shHMGA1) and treated for 6 days with 4OHT before ATAC-seq.