RAD21 is a subunit of cohesin. Cohesin is maninly known for its role in holding sister chromatids together during cell-division but it also plays a role in gene regulation. Genome wide study in MCF7 cells have shown that cohesin co-binds with estrogen receptor alpha at numerous sites. To identify estrogen sensitive genes that are influenced by cohesin, a microarray expression analysis was conducted in MCF7 cells lacking the cohesin subunit RAD21. Global changes in gene expression were identified in cells treated with Control/RAD21 siRNA and stimulated with estrogen/vehicle thereafter.
Overall design
MCF7 cells growing in hormone depleted conditions for 24 hours were transfected with RAD21 or control siRNA and cultured in hormone-depleted conditions for a further 48 hours. Cells were subsequently treated with 100 nM 17-β estradiol or vehicle for 3 and 6 hours. Cells were harvested for RNA and protein at both time points. Knockdown of RAD21 message and protein were verified by quantitative PCR and western blotting respectively. For each time point, RNA from three independent biological experiments were used for analyses
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