Mechanisms Whereby Src Activates Estrogen Stimulated ER Proteolysis and ER Target

Src 激活雌激素刺激的 ER 蛋白水解和 ER 靶标的机制

基本信息

批准号：
7799932
负责人：
JOYCE MARIE SLINGERLAND
金额：
$ 33.29万
依托单位：
UNIVERSITY OF MIAMI SCHOOL OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-04-06 至 2014-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7799932
关键词：
26S proteasome Affect Aggressive Clinical Course Animals Back Binding Biological Assay Breast Cause of Death Chromatin Coupled Data Epidermal Growth Factor Receptor Estrogen Receptors Estrogen receptor positive Estrogens Feedback Feeds Gene Expression Gene Targeting Genetic Genetic Transcription Grant Human In Vitro Label Ligands Link Malignant Neoplasms Mammary gland Mediating Messenger RNA Methionine Microarray Analysis Molecular Target Mus Oncogenic Phosphorylation Proteasome Inhibition Proteasome Inhibitor Protein Biosynthesis Proteins Proteolysis Receptor Activation Receptor Protein-Tyrosine Kinases Recruitment Activity Regulation Resistance Series Signal Transduction Testing Time Tissues Transcriptional Activation Transgenic Organisms Tyrosine Phosphorylation Ubiquitin Ubiquitin-Activating Enzymes Ubiquitin-Conjugating Enzymes Woman Work activating transcription factor base cancer type deprivation in vivo inhibitor/antagonist malignant breast neoplasm new therapeutic target non-genomic outcome forecast overexpression prevent prognostic public health relevance src-Family Kinases steroid hormone transcription factor tumor ubiquitin ligase

项目摘要

DESCRIPTION (provided by applicant): Breast cancer is the most frequent womens' cancer. One third of new breast cancers are estrogen receptor a (ER) protein negative and have a worse prognosis than ER positive breast cancers. The ER is a ligand activated transcription factor. Estrogen:ER binding stimulates rapid Src activation that feeds back to phosphorylate ER and increases its transcriptional activity. Estrogen rapidly activates ubiquitin-dependent ER proteolysis which in turn regulates ER activity. Our data suggest that Src-stimulates ER proteolysis and this is linked to activation of certain ER target genes. Src induction increased both ER-activated gene expression and ER proteolysis. Src inhibition increased ER stability. The weakly ER +, MDA-MB-361 and ER negative, BT-20 breast cancer lines both have high Src, and while ER protein synthesis was easily detected, the ER t1/2 was reduced. ER was increased by both estrogen deprivation and proteasome inhibitors in both ER + and ER- line studied. SiRNA to Src increased ER protein in BT-20. E6AP is an ubiquitin ligase that also acts as an ER- coactivator. We show E6AP acts as ubiquitin ligase for ER in vitro and E6AP-mediated ER ubiquitylation was increased by ER pre-treatment with Src kinase. Src inhibitors impaired ubiquitylation and degradation of ER in vivo and in vitro. Src activity was increased in primary ER negative breast cancers compared to ER positive. We further investigate if Src, when recruited by activated ER, regulates transcription-coupled ER proteolysis. Our Hypothesis is that a subset of ER negative breast cancers are estrogen responsive: they express ER mRNA but ER protein is undetectable due to accelerated Src and E6AP-mediated ER proteolysis. Oncogenic Src activation may phosphorylate the ER or key co-regulators to activate both ER proteolysis and ER target gene transcription. This is pursued in Aims 1-3: 1. To further test how Src activation affects ER levels in primary human breast cancers and in breast cancer lines; 2. To test whether Src mediated phosphorylation of the ER or of a key co-activator can promote ER ubiquitylation by E6AP and ER degradation in vitro and in vivo; and 3. To assay if Src activation modulates ER transcriptional activity. Oncogenic Src promotes breast cancer proliferation and survival, and may also accelerate ER proteolysis in breast cancers. Elucidation of mechanisms underlying the ER negative status of breast cancer may indicate why they are so aggressive and yield new therapeutic targets for this treatment-resistant breast cancer type. PUBLIC HEALTH RELEVANCE: One third of new breast cancers are estrogen receptor a (ER) protein negative and have a worse prognosis than ER positive breast cancers. This grant pursues the hypothesis that a subset of ER negative breast cancers are estrogen responsive: they express ER mRNA but ER protein is undetectable due to accelerated Src and E6AP-mediated ER proteolysis. Oncogenic Src activation may phosphorylate the ER or key co- regulators to activate both ER proteolysis and ER target gene transcription. Oncogenic Src promotes breast cancer proliferation and survival. Our work suggests that it may also accelerate ER proteolysis in breast cancers. Elucidation of mechanisms underlying the ER negative status of breast cancer may indicate why they are so aggressive and yield new therapeutic targets for this treatment-resistant breast cancer type.

描述（由申请人提供）：乳腺癌是最常见的女性癌症。三分之一的新乳腺癌是雌激素受体A（ER）蛋白阴性的，并且预后较差。 ER是配体激活的转录因子。雌激素：ER结合刺激快速的SRC激活，从而恢复磷酸化ER并增加其转录活性。雌激素迅速激活泛素依赖性的ER蛋白水解，从而调节ER活性。我们的数据表明SRC刺激ER蛋白水解，这与某些ER靶基因的激活有关。 SRC诱导增加了ER激活的基因表达和ER蛋白水解。 SRC抑制增加了ER稳定性。弱的ER +，MDA-MB-361和ER阴性，BT-20乳腺癌系都有高SRC，尽管很容易检测到ER蛋白的合成，但ER T1/2却降低了。 ER +和ER线研究中的雌激素剥夺和蛋白酶体抑制剂都增加了ER。 siRNA至SRC增加了BT-20中的ER蛋白。 E6AP是一种泛素连接酶，也充当ER-共激活剂。我们表明，通过用SRC激酶进行ER预处理，增加了体外ER和E6AP介导的ER泛素化的泛素连接酶的作用。 SRC抑制剂损害了体内和体外ER的泛素化和降解。与ER阳性相比，原发性ER负乳腺癌的SRC活性增加。我们进一步研究SRC是否通过激活的ER募集时，会调节转录耦合的ER蛋白水解。我们的假设是，ER负乳腺癌的子集具有雌激素的反应：它们表达了ER mRNA，但由于加速的SRC和E6AP介导的ER蛋白：无法检测到ER蛋白。致癌性SRC激活可能会磷酸化ER或关键共同调节剂，以激活ER蛋白水解和ER靶基因转录。在目标1-3：1中提出了这一点，以进一步测试SRC激活如何影响原发性人乳腺癌和乳腺癌系中的ER水平； 2。为了测试SRC介导的ER或关键共激活因子的磷酸化是否可以在体外和体内通过E6AP和ER降解来促进ER泛素化； 3。如果SRC激活调节ER转录活性，则测定。致癌性SRC促进乳腺癌的增殖和存活，也可能加速乳腺癌的ER蛋白水解。阐明乳腺癌负面状态负面状态的机制可能表明为什么它们如此侵略性并为这种耐药性乳腺癌类型产生新的治疗靶标。公共卫生相关性：三分之一的新乳腺癌是雌激素受体A（ER）蛋白质阴性的，并且预后较差，而ER乳房癌的预后较差。该赠款提出了以下假设：ER负乳腺癌的子集具有雌激素的反应：它们表达了ER mRNA，但由于加速的SRC和E6AP介导的ER蛋白水解而无法检测到ER蛋白。致癌性SRC激活可能会磷酸化ER或关键共同调节剂，以激活ER蛋白水解和ER靶基因转录。致癌性SRC促进乳腺癌的增殖和存活。我们的工作表明，它也可能加速乳腺癌中的ER蛋白水解。阐明乳腺癌负面状态负面状态的机制可能表明为什么它们如此侵略性并为这种耐药性乳腺癌类型产生新的治疗靶标。