A novel serine phosphorylation site detected in the n-terminal domain of estrogen receptor isolated from human breast cancer cells

Britton, David; Scott, Gary K.; Schilling, Birgit; Atsriku, Christian; Held, Jason M.; Gibson, Bradford W.; Benz, Christopher C.; Baldwin, Michael A.

doi:10.1016/j.jasms.2008.02.008

Cited by 28 publications

(26 citation statements)

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“…More recently we enriched and immunoaffinity-purified endogenous ER␣ derived from the human breast cancer cell line MCF-7 in quantities compatible with tandem MS analysis and identified a novel phosphorylation site, Ser 154 . vMALDI-MS n confirmed Ser 154 phosphorylation in human breast cancer cells grown under both ligand-dependent and ligand-independent conditions (27). Coincidentally we also observed known phosphorylation sites Ser 118 and Ser 167 not previously confirmed by MS methods.…”

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confidence: 76%

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Systematic Mapping of Posttranslational Modifications in Human Estrogen Receptor-α with Emphasis on Novel Phosphorylation Sites

Atsriku

Britton

Held

et al. 2009

Molecular & Cellular Proteomics

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A systematic study of posttranslational modifications of the estrogen receptor isolated from the MCF-7 human breast cancer cell line is reported. Proteolysis with multiple enzymes, mass spectrometry, and tandem mass spectrometry achieved very high sequence coverage for the full-length 66-kDa endogenous protein from estradiol-treated cell cultures. Nine phosphorylated serine residues were identified, three of which were previously unreported and none of which were previously observed by mass spectrometry by any other laboratory. Two additional modified serine residues were identified in recombinant protein, one previously reported but not observed here in endogenous protein and the other previously unknown. Although major emphasis was placed on identifying new phosphorylation sites, N-terminal loss of methionine accompanied by amino acetylation and a lysine side chain acetylation (or possibly trimethylation) were also detected. The use of both HPLC-ESI and MALDI interfaced to different mass analyzers gave higher sequence coverage and identified more sites than could be achieved by either method alone. The estrogen receptor is critical in the development and progression of breast cancer. One previously unreported phosphorylation site identified here was shown to be strongly dependent on estradiol, confirming its potential significance to breast cancer. Greater knowledge of this array of posttranslational modifications of estrogen receptor, particularly phosphorylation, will increase our understanding of the processes that lead to estradiol-induced activation of this protein and may aid the development of therapeutic strategies for management of hormonedependent breast cancer. Molecular & Cellular Proteomics 8:467-480, 2009.The ␣-isoform of the estrogen receptor (ER␣) 1 is a 66-kDa nuclear transcription factor that mediates transcriptional regulation of genes involved in cell proliferation and differentiation and plays a pivotal role in the development and progression of breast cancer (1-3). Consequently the development of therapies for management of hormone-dependent breast cancer has targeted signaling pathways based on modulation of ER␣ activity (4 -6). Current knowledge and understanding of ER␣ activity is derived from over 30 years of accumulated scientific evidence that has sought to delineate ER-mediated mechanisms and signaling pathways under pathological conditions modeled in cultured breast cancer cell lines. Because the function or activity of a protein may depend strongly on the presence of posttranslational modifications (PTMs), significant research has focused on detection and quantitation of such modifications in ER␣. The constellation of PTMs on a protein constitutes a molecular code that may dictate protein conformation, localization, and function. Thus biological inference based on ER␣ structure requires a comprehensive study of all possible PTMs on the constituent amino acids.Modifications to ER␣ reported to date are listed in Table I. Two of the most common and important PTMs that modulate the...

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confidence: 76%

“…Reagents and Chemicals-These were as described previously (27) except that human recombinant ER␣ (rER␣) was from Invitrogen, and sequencing grade chymotrypsin, Lys-C, Asp-N, and Glu-C were from Roche Diagnostics.…”

Section: Methodsmentioning

confidence: 99%

Systematic Mapping of Posttranslational Modifications in Human Estrogen Receptor-α with Emphasis on Novel Phosphorylation Sites

Atsriku

Britton

Held

et al. 2009

Molecular & Cellular Proteomics

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“…1 et al 1995b). Recently, novel phosphorylation sites in ERa were identified (Britton et al 2008. Table 1 lists the sites of phosphorylation in ERa, which have been identified experimentally, using different methodologies.…”

Section: Phosphorylation Sites Identified In Eramentioning

confidence: 99%

“…The presence of multiple phosphorylation sites in ERa (Britton et al 2008, Murphy et al 2009a) that may have differential effects on activity suggests that it may be necessary to consider the balance of multiple phosphorylation sites in vivo in terms of predicting clinical outcome with respect to endocrine treatment responsiveness. Recently, data have been published where up to seven different phosphorylation sites in ERa in any individual tumor were taken into account by developing a mathematical model that balances the presence of phosphorylation sites associated with good clinical benefit and those associated with poor clinical benefit.…”

Section: Multiple Phosphorylated Forms Of Eramentioning

confidence: 99%

Clinical significance of estrogen receptor phosphorylation

Murphy

Seekallu²,

Watson³

2010

Endocrine Related Cancer

128

106

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Multiple sites of phosphorylation on human estrogen receptor a (ERa) have been identified by a variety of methodologies. Now with the emerging availability of phospho-site-specific antibodies to ERa, the relevance of phosphorylation of ERa in human breast cancer in vivo is being explored. Multiple phosphorylated sites in ERa can be detected in multiple breast tumor biopsy samples, providing evidence of their relevance to human breast cancer in vivo. Published data suggest that the detection in primary breast tumors of phosphorylation at some sites in ERa is associated with a better clinical outcome while phosphorylation at other sites is associated with a poorer clinical outcome most often in patients who have been treated with tamoxifen. This suggests the hypothesis that phospho-profiling of ERa in human breast tumors to establish an 'ERa phosphorylation code', may be a more accurate marker of prognosis and/or response to endocrine therapy in human breast cancer.

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“…This EGF-induced ER escape from hormone regulation could be explained by evidence that EGF triggers ligand-independent ER transcriptional activity through phosphorylation of specific critical residue(s) of nuclear estrogen receptor promoting interaction with its co-activator SRC1 (Schiff et al, 2003). EGF stimulates through MAPK-dependent signaling ERalpha serine phosphorylation on 118 (Kato et al, 1995) and 154 residues (Britton et al, 2008), likely enhances 305 serine phosphorylation (Rayala et al, 2006), and induces 167 serine phosphorylation through a PI3-K dependent pathway (Campbell et al, 2001). It has also been shown that EGF activated MAPK pathway can phosphorylate the ER coactivator AIB1 and nuclear co-repressor NCoR (Font de Mora & Brown, 2000).…”

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confidence: 99%

Cross talk between epidermal growth factor (EGF) receptor and extra nuclear steroid receptors in cell lines

Migliaccio

Castoria

Giovannelli

et al. 2010

Molecular and Cellular Endocrinology

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Please cite this article as: Migliaccio, A., Castoria, G., Giovannelli, P., Auricchio, F., cross talk between epidermal growth factor (egf) receptor and extranuclear steroid receptors in cell lines, Molecular and Cellular Endocrinology (2010), doi:10.1016/j.mce.2010 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. Page 1 of 19A c c e p t e d M a n u s c r i p t CROSS TALK BETWEEN EPIDERMAL GROWTH FACTOR (EGF) RECEPTOR AND EXTRANUCLEAR STEROID RECEPTORS IN CELL LINES.Antimo Migliaccio, Gabriella Castoria, Pia Giovannelli and Ferdinando Auricchio. Department of General Pathology-II Università di Napoli Via L. De Crecchio, 7 80138 Napoli (ITALY)Key words: EGF, EGF-R, steroid receptors, cross talk, Src-dependent signaling pathway.Email: antimo.migliaccio@unina2.it *ManuscriptPage 2 of 19 A c c e p t e d M a n u s c r i p t 2 ABSTRACTSteroid receptors act as ligand-dependent transcriptional factors. It has been observed that in addition to responding to cognate hormones with transcription activation, once hormone bound they are also capable of rapid responses following association with signaling effectors in the extra nuclear compartment. This novel aspect of steroid hormone action could influence our view of the cross talk between growth factor and steroid receptors. Increasing evidence shows that in hormone-responsive cells, a cross talk occurs between growth factors (EGF, IGF-1) and steroid hormone receptors that reciprocally regulate their action. To date, this has mostly been explained by modulation of steroid receptor transcriptional activity through growth factor receptor signaling activation. However, it is now known that growth factors might also act on extra nuclear steroid receptors, activating them via a hormone-independent mechanism. On the other hand, extranuclear steroid receptors can regulate growth factor receptor activity either directly interfering with their transduction pathways, or inducing autocrine growth factor secretion. Here we discuss findings indicating that EGF, like steroid hormones, induces association of steroid receptors with Src thereby activating pathways that can trigger cell proliferation and migration. Since mammary and prostate cancers respond to both steroid hormones and growth factors, this association might be a putative target for human cancer therapy. Findings from our laboratory supporting this view are discussed. Page 3 of 19A c c e p t e d M a n u s c r i p t 3 THE COMPLEX NETWORK OF EXTRA NUCLEAR INTERACTIONS OF STEROID RECEPTORS: THE EXAMPLE OF ESTROGEN RECEPTOR ALPHAEstrogen receptor/s (ERs) interact with a multitude of proteins inside as well as outside the...

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A novel serine phosphorylation site detected in the n-terminal domain of estrogen receptor isolated from human breast cancer cells

Cited by 28 publications

References 50 publications

Systematic Mapping of Posttranslational Modifications in Human Estrogen Receptor-α with Emphasis on Novel Phosphorylation Sites

Systematic Mapping of Posttranslational Modifications in Human Estrogen Receptor-α with Emphasis on Novel Phosphorylation Sites

Clinical significance of estrogen receptor phosphorylation

Cross talk between epidermal growth factor (EGF) receptor and extra nuclear steroid receptors in cell lines

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