Extranuclear signaling by sex steroid receptors and clinical implications in breast cancer

Abstract

Estrogen and progesterone play essential roles in the development and progression of breast cancer. Over 70% of breast cancers express estrogen receptors (ER) and progesterone receptors (PR), emphasizing the need for better understanding of ER and PR signaling. ER and PR are traditionally viewed as transcription factors that directly bind DNA to regulate gene networks. In addition to nuclear signaling, ER and PR mediate hormone-induced, rapid extranuclear signaling at the cell membrane or in the cytoplasm which triggers downstream signaling to regulate rapid or extended cellular responses. Specialized membrane and cytoplasmic proteins may also initiate hormone-induced extranuclear signaling. Rapid extranuclear signaling converges with its nuclear counterpart to amplify ER/PR transcription and specify gene regulatory networks. This review summarizes current understanding and updates on ER and PR extranuclear signaling. Further investigation of ER/PR extranuclear signaling may lead to development of novel targeted therapeutics for breast cancer management.

Keywords: Breast cancer; Estrogen receptor; Growth factor signaling; Nongenomic signaling; Progesterone receptor; Rapid membrane signaling.

Figure 1. Estrogen receptor signaling pathways in human breast tumors

Proliferation and survival of BC cells is closely regulated by estradiol-17β (E2) and its receptors, ERα and ERβ, as well as growth factor receptors. In classic models of E2 action (1), E2 binds ER to promote dimerization and phosphorylation of ER. This allows direct binding of the E2-ER complex with steroid receptor coactivators and estrogen response elements (ERE) in DNA, leading to gene transcription to regulate growth, differentiation, apoptosis and angiogenesis. A subset of ERs occur in extranuclear sites, such as caveolae or lipid rafts in plasma membrane (2), and may interact with transmembrane growth factor receptors such as EGFR (3), HER2 (4), insulin-like growth factor receptor I (IGFR1) (5) and other signaling molecules, including components of Ras-MAPK and phosphatidylinositol 3-kinase (PI3K)/AKT pathways, Src kinases, Janus-activated kinase/signal transducer and activator of transcription signaling, nitric oxide synthase (NOS) (6), and G-proteins (7). Membrane-associated ER may undergo posttranslational modification, such as palmitoylation, and/or associate with adaptor proteins, such as Shc, PELP1, or lipid raft proteins. ERs and growth factor receptors may form a structured complex for signal transduction to MAPK and/or PI3K/AKT kinase that interacts, in turn, with nuclear ER and steroid receptor coactivators (8). Signaling for cell growth involves phosphorylation (P) of nuclear ER and coactivators that can occur in ligand-dependent as well as ligand-independent modes. ERE-dependent and alternate transcription sites may be activated (9). Further, E2 is produced locally in BC and in host supporting cells by the action of aromatase (ARO) (10) which is regulated by both nuclear and extranuclear ER. Pathways modified from Pietras and Marquez [52]; refer to text for more details.

Figure 2. Supramolecular organization of plasma membrane and occurrence of estrogen receptors (ER)

A model of the surface membrane from an estradiol (E2)-responsive cell in the region of a caveolar structure is shown. E2 may interact with one of several different forms of membrane-associated ERs. Full structural characterization of these ERs remain to be done. These molecules may be known membrane components, such as enzymes, G-proteins, ion channels, or receptors for nonsteroidal ligands, with unrecognized steroid binding sites (1); new isoforms of hormone receptors such as the truncated ERs that arise by alternative splicing (2); ‘classical’ receptors complexed with other membrane-associated proteins (3); or novel membrane proteins (4). Of note, splice variants of ER occur, and these give rise to proteins of different molecular size and possibly modified properties. Membrane insertion of receptors in primary transcript form would likely require one or more hydrophobic regions. ERα, for example, has several hydrophobic regions, but it is not known if these suffice for disposition as integral membrane proteins. Posttranslational modification of receptor protein leading to membrane targeting also occurs, including phosphorylation, glycosylation, nitrosylation and/or addition of lipid anchors or alterations such as palmitoylation or myristoylation. Evidence for palmitoylation of ER that fosters membrane association is documented in the text. Modified from Szego and Pietras [322].

Figure 3. Convergence of PR nuclear and extranuclear signaling pathways to regulate biological responses

In the classic nuclear signaling pathway (1) progesterone activates progesterone receptor (PR) by binding and inducing conformational changes of the receptor causing in turn nuclear translocation, dimerization and binding to progesterone response elements (PRE) in the promoters or enhancer regions of PR target genes. Progestin treatment rapidly activates extranuclear signaling of a subpopulation of PR localized in the membrane or cytoplasm to transiently associate with c-Src through interaction with the PR polyproline domain (PPD) leading to activation of mitogen-activated protein kinase (MAPK), phosphatidylinositol 3-kinase (PI3K/Akt), or Janus kinase/signal transducers and activators of transcription (JAK/STAT) signaling pathways. Activated MAPK further regulates PR transcriptional activity through phosphorylation of PR, co-activators (1) or other transcription factors (2). Rapid progestin activation of MAPK can directly phosphorylate PR or co-activators (1) or promote PR phosphorylations through CK2 and CDK2 (see text for details). Alternatively, activation of various cytoplasmic signaling pathways may phosphorylate and increase transcriptional activity of other transcription factors, independent of PREs (2). Rapid extranuclear activation of ER/PR complexes by progestins activates MAPK leading to a formation of phospho-PR/MAPK/Msk1 (Mitogen and stress-activated protein kinase 1), chromatin remodeling and enhanced MMTV and PRE-containing gene transcription (3). PR extranuclear activation of Src/PI3K/Akt may stimulate focal adhesion kinase (FAK) or ribosomal S6 kinase 1 (RSK1) triggering actin cytoskeleton remodeling and promotion of cell motility (4). Alternatively, PR extranuclear signaling may cross-communicate with growth factor signaling such as epidermal growth factor (EGF) leading to activation of MAPK and downstream events (5). Membrane localized progestin receptor (mPR) unrelated to the classical PR mediates progestin extranuclear signaling through GPCR-like membrane proteins via modulation of adenylate cyclase (AC), cyclic adenosine monophosphate (cAMP) and protein kinase A (PKA). mPR has been reported to activate MAPK and PI3K/Akt signaling pathways. More work is needed to define the physiological significance of extranuclear mPR activation of these several cytoplasmic signaling pathways in BC cells.