MECHANISMS OF SIGNAL TRANSDUCTION
Novel Activation of STAT5b in Response to Epidermal Growth Factor*

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Of the seven signal transducers and activators of transcription that have been identified, STATs 1, 3, and 5a/5b can be activated not only by a multitude of cytokines but also by some growth factors. The data presented here demonstrate that, in contrast to activation by the cytokine, growth hormone (GH), the activation of STAT5b by the growth factor, epidermal growth factor (EGF), requires overexpression of the EGF receptor (EGFR). We have shown that EGF activates STAT5b not only in a HEK293 cell model in which the EGFR is stably overexpressed but also in the MDA-MB468 breast cancer cell line. Furthermore, EGF (but not GH) is able to activate tyrosine phosphorylation of a Tyr-699 mutant of STAT5b. Using metabolic labeling studies as well as site-directed mutagenesis, we have identified three novel EGF-induced tyrosine phosphorylation sites, Tyr-725, Tyr-740, and Tyr-743. Luciferase assays using a STAT5-specific DNA sequence demonstrate that, although Tyr-699 is absolutely required for transcriptional activation, tyrosines 725, 740, and 743 may be involved in a negative regulation of transcription. Because overexpression of the EGFR is common in many cancers, including advanced breast cancer, characterization of EGF-induced STAT5b may have direct implications in therapeutic applications.

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Published, JBC Papers in Press, December 20, 2001, DOI 10.1074/jbc.M111884200

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This work supported by a Career Development Award from the Department of Defense (DAMD17-99-1-9431) and Grant R01-CA85462 from NCI/National Institutes of Health (NIH) (to C. M. S.). Construction of the Y699F mutant was done by the Molecular Biology Core Laboratory at the University of Virginia, which was supported by NICHD/NIH through, cooperative agreement U54 HD28934 as part of the Specialized Cooperative Centers Program in Reproduction Research.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.