Rho GTPases as Modulators of the Estrogen Receptor Transcriptional Response

Abstract

The estrogen receptor ␣ (ER) is a ligand-dependent transcription factor that plays a critical role in the development and progression of breast cancer, in part, by regulating target genes involved in cellular proliferation. To identify novel components that affect the ER transcriptional response, we performed a genetic screen in yeast and identified RDI1, a Rho guanine nucleotide dissociation inhibitor (Rho GDI), as a positive regulator of ER transactivation. Overexpression of the human homologue of RDI1, Rho GDI␣, increases ER␣, ER␤, androgen receptor, and glucocorticoid receptor transcriptional activation in mammalian cells but not activation by the unrelated transcription factors serum response factor and Sp1. In contrast, expression of constitutively active forms of RhoA, Rac1, and Cdc42 decrease ER transcriptional activity, suggesting that Rho GDI increases ER transactivation by antagonizing Rho function. Inhibition of RhoA by expression of either the Clostridium botulinum C3 transferase or a dominant negative RhoA resulted in enhanced ER transcriptional activation, thus phenocopying the effect of Rho GDI expression on ER transactivation. Together, these findings establish the Rho GTPases as important modulators of ER transcriptional activation. Since Rho GTPases regulate actin polymerization, our findings suggest a link between the major regulators of cellular architecture and steroid receptor transcriptional response. The estrogen receptor ␣ (ER)1 is a ligand-dependent transcription factor that transduces the estrogen signal (1). Activation of ER is responsible for female sexual development and maintenance of bone density (2, 3). In addition, ER plays a critical role in the development and progression of breast cancer by regulating genes and signaling pathways involved in cellular proliferation (4). Regulation of gene expression by the ER requires the coordinate activity of ligand binding, phosphorylation, and cofactor interactions, with particular combinations probably resulting in the tissue-specific responses elicited by the receptor (5-7). However, the extracellular cues and intracellular signaling pathways modulating these components and regulating ER transcriptional activation are not fully understood.To identify novel proteins that modulate ER transcriptional activation, we have carried out a genetic screen in the yeast Saccharomyces cerevisiae. The ability of the human ER to function within yeast allows a wide variety of genetic approaches to be taken toward further defining the mechanism of ER transcriptional activation given the ease of genetic manipulation and simplicity of gene identification in yeast. In addition, with the large number of orthologous proteins carrying out the same biological functions in both S. cerevisiae and metazoans (8 -10), it is likely that the yeast factors affecting ER transactivation will have mammalian counterparts, which can be examined in vertebrate systems.The genetic approach we have used to identify factors that affect ER transcriptional activation is dosage su...

Rho GDP dissociation inhibitor α interacts with estrogen receptor α and influences estrogen responsiveness

“…4). These findings are in agreement with previous transfection experiments carried out in U2OS cells (Su et al 2001).…”

“…Likewise, the localization of RhoGDIa in the nucleolus may help to simultaneously sequester RhoGDIa from cytoplasmic GTPases and from ERa and at the same time serve as a storage depot to maintain a pool of nuclear RhoGDIa protein that could interact with ERa and influence gene expression. The ability of RhoGDIa to alter ERa activity was previously reported by Garabedian and coworkers (Su et al 2001), who showed that RhoGDIa enhances ERa, ERb, glucocorticoid receptor and androgen receptor (AR), but not SRF or Sp1, mediated transactivation in transient transfection assays. A subsequent study by this group suggested that RhoGDIa enhances ERamediated transactivation indirectly through its effects on ERa-associated coregulatory proteins (Su et al 2002).…”

“…One of these ERa-associated proteins was the 28 kDa protein Rho GDP dissociation inhibitor a (RhoGDIa), which was originally characterized as a negative regulator of the RhoGTPase family members RhoA, Rac1, and Cdc42 (Fukumoto et al 1990, Leonard et al 1992, Masuda et al 1994, Koch et al 1997, Olofsson 1999. Although the ability of RhoGDIa to enhance transcription of an estrogen response element (ERE)-containing reporter plasmid has been reported previously, it was thought that this enhanced activity was due to the cytoplasmic actions of RhoGDIa (Su et al 2001(Su et al , 2002). Because we found RhoGDIa associated with the DNA-bound ERa, we investigated the localization of endogenously expressed RhoGDIa in MCF-7 breast cancer cells and characterized the ability of this protein to interact with ERa and influence the expression of estrogen-responsive genes.…”

Energetic determinants of protein binding specificity: Insights into protein interaction networks

“…This protein contains a promiscuous binding site that mediates two different interactions, its regulator rho-GDI (rdi1) [50], and the GTPase-activating protein lrg1 [51]. This interface comprises four modules (Fig.…”

Distinct Transcriptional Regulation of Human Large Conductance Voltage- and Calcium-activated K+ Channel Gene (hSlo1) by Activated Estrogen Receptor α and c-Src Tyrosine Kinase

“…Described signaling cascades resulting in transcriptional activation are as follows: c-Src tyrosine kinase and downstream MAPK, PI3K, and mitogen/ERK kinase-1 (15)(16)(17)(18)(19). Also, monomeric G-protein Rho pathways are involved in ER␣-mediated transcriptional activity * This work was supported, in whole or in part, by National Institutes of Health (20). In the case of mSlo1, a genomic mechanism defines the up-regulation by estrogen-activated ER␣, where ER␣ binds to quasi-perfect EREs in the mSlo1 promoter, a mechanism that may be complemented by Sp1 transcription factor (21).…”