Tools to study the function of the Ras-related, estrogen-regulated growth inhibitor in breast cancer

The Ras-related, estrogen-regulated growth inhibitor (Rerg) is a Ras-related small GTPase and candidate tumor suppressor. Rerg gene expression is stimulated by the estrogen receptor alpha (ER alpha), and Rerg gene expression is absent in ER-negative breast cancers. ER-negative breast cancers are highly invasive and metastastic and are typically more advanced than their ER-positive counterparts. Like Ras, Rerg binds and hydrolyzes GTP, but unlike Ras, Rerg has been shown to possess growth inhibitory activity in breast cancer cells. The precise role that Rerg loss plays in breast cancer growth and the mechanisms by which it does so are unknown. This chapter describes tools used to detect and manipulate the expression of Rerg in breast cancer cells. We validate use of an antibody to detect Rerg expression. We describe the generation of expression vectors that encode wild-type and mutants of Rerg that are altered in GDP/GTP regulation. We also describe the development of an inducible Rerg expression system and of a retrovirus- based RNA interference approach to repress Rerg expression. These toots will be invaluable in evaluating the biological function of Rerg in breast cancer.