Systematic mapping of posttranslational modifications in human estrogen receptor-α with emphasis on novel phosphorylation sites

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

Research output: Contribution to journalArticlepeer-review

58 Scopus citations

Abstract

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 hormone-dependent breast cancer.

Original languageEnglish
Pages (from-to)467-480
Number of pages14
JournalMolecular and Cellular Proteomics
Volume8
Issue number3
DOIs
StatePublished - Mar 2009

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