During FY2001 we built upon key observations that we made concerning the mechanisms by which steroid receptors activate transcription within breast cancer cells. We demonstrated that proteins involved in the manipulation of chromatin structure, chromatin remodeling machines, represent the key components in the cascade of events that results in the activation of the genetic program in human breast cancer cells. We have made significant advances in understanding the nature of the cellular response to prolonged glucocorticoid exposure. Our studies revealed that there is a profound reduction in the level of phosphorylated histone H1 that is directly linked to the cessation of transcription. The mechanism that underpins this result is the inhibition of the key cell cycle regulator CDK2 in response to glucocorticoid. In congruence with our hypothesis that understanding chromatin structure is vital to understanding gene regulation these events take place only in the context of chromatin. We continued to analyze the activity of the glucocorticoid receptor within breast and osteosarcoma cancer cells that differ in the expression of components of the chromatin remodeling machines. These cells display an altered response to a variety of clinically important hormone antagonist and will be useful in evaluating various anti-hormone strategies in breast cancer. In addition they are a unique resource to evaluate and characterize the impact of a number of environmental agents in human cells. To this end we have continued to develop methodologies that allow us to look at protein-DNA interactions within living cells, in vivo footprinting, as well as changes at the control regions of specific genes within living cell, chromatin immunoprecipitation (CHIP) assays.
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