2005-2010

Our long term goal is to determine the mechanism(s) by which the nuclear receptor coactivator AIB1 (amplified in breast cancer 1) potentiates growth factor signaling. AIB1 (and its more active isoform AIB1-?3) is the only member of the SRC (steroid receptor coactivator) family that is amplified and overexpressed in a wide variety of epithelial tumors suggesting a major role in neoplastic development. In the past 3 years we have reported that: a) overexpression of AIB1-?3 in transgenic mice causes pronounced mammary hyperplasia and changes in levels of proliferative and apoptotic signaling molecules b) AIB1 is rate limiting for estrogen (E) and growth factor induced proliferation, anti-apoptosis and in vivo tumor growth of breast cancer cell lines. In addition to this role of AIB1 in hormone signaling, emerging data also show an role for AIB1 in the development of the hormone-independent human breast cancer: a) We were the first to report that AIB1 and AIB1-3 can potentiate EGF induction of transcription. b) We also found that AIB1 is rate limiting for EGF and heregulin- induced phenotypic changes important to tumor progression in vitro. c) We have determined that reductions in AIB1 reduce IGF-1 (insulin-like growth factor 1) inhibition of anoikis and stimulation of proliferation. Together, these data indicate a central role for AIB1 in tyrosine kinase receptor signaling pathways that are of major importance for the development of breast cancer. The mechanism(s) of AIB1 potentiation of IGF-1R and EGFR family signaling is (are) not known and will be studied here. We have reported that AIB1 is detected in the cytosol of normal breast epithelium and changes its location to the nuclei in breast cancer epithelia. In cultured cells we observed a rapid increase in tyrosine phosphorylation of AIB1 in the cytoplasm within minutes of IGF-1, EGF or heregulin treatment. This is paralleled by nuclear translocation of AIB. These effects are mimicked and potentiated by the phosphatase inhibitor, orthovanadate. Finally, we have observed that IGF-1 promotes the rapid association of AIB1 with several critical cytoplasmic signaling molecules, most notably IRS-1. To our knowledge this is the first report of regulatable tyrosine phosphorylation of a coactivator and the first report of the regulatable interaction of a coactivator with the cytoplasmic signaling molecule IRS-1. Based on these studies, we hypothesize that tyrosine phosphorylation of AIB1 promotes interactions with critical signaling molecules in the IGF-1R and EGFR family pathways. These events initiate the nuclear translocation and transcriptional coactivator activity of AIB1. The hypothesized steps are indicated in the model shown to the right. Our hypotheses will be addressed in two Specific Aims: Aim 1: To determine the mechanisms and the function of tyrosine phosphorylation of AIB1 due to IGF-1, EGF and Heregulin stimulation of cells. A) we will identify the tyrosine residues phosphorylated in AIB1 after IGF, EGF or Heregulin treatment B) we will determine the role of these tyrosine residues in AIB1 for: (i) translocation of AIB1 to the nucleus (ii) transcriptional activity of AIB1 C) we will determine the upstream pathways leading to tyrosine phosphorylation of AIB1 due to IGF-1, EGF and Heregulin stimulation. Aim 2: To determine the role of AIB1 in growth factor mediated mammary tumorigenesis. A) we will determine whether gain or loss of AIB1 or AIB1-?3 alters preneoplasia, onset, incidence, metastatic spread and gene expression pattern of mammary adenocarcinoma in MMTV/neuT mice B) we will determine if AIB1 directly effects IGF-1, EGF or Heregulin pathways by examining signaling in mammary organ culture derived from MMTV/neuT mice with gain or loss of AIB1 or AIB1-?3.

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