Breast cancer cells can recognize and respond to different levels of progestins to achieve different phenotypic outputs [ATAC-seq]
The steroid hormone progesterone, acting through its nuclear progesterone receptor (PR), has complex physiologic activities with different levels of hormones manifesting distinct and sometimes opposing phenotypic responses in target tissues. However, most of what is currently known about the transcriptional activity of PR comes from studies performed using progestins at levels that are in the high physiologic range (>10nM), relevant only in the luteal phase of the reproductive cycle and pregnancy in humans. These studies do not consider the non-linearity of responses to progestins that exist in physiology and are not informative as to the mechanisms by which low levels of progestins, as occurs during menopause (0.1-0.3nM), exert their biological activities. Thus, we undertook to define the mechanisms which enable cells to recognize and respond to different levels of progestins. Overall design: T47D cells were plated 2x10^5 cells per well in 10cm culture plates. Cells were allowed to attach overnight in phenol red-free RPMI 1640 supplemented with 8% charcoal-stripped FBS (CFS), NEAA, and NaPyr. Synchronization was achieved by 48hrs serum starvation in phenol red-free RPMI 1640 supplemented with 0.1% CFS, NEAA, and NaPyr. Synchronized T47D cells were then treated with 0.1nM R5020, 10nM R5020, or DMSO vehicle. Cell plates were incubated at 37*C in the presence of 5% CO2 for 6hrs or 18hrs, after which cells were washed with PBS dissociated from the culture plate using gentle trypsinization (0.05% for minimal time). Omni-ATAC was performed as previously described (PMID: 28846090).