Rho GEFs in Cellular Proliferation and Transformation

Rho GEF 在细胞增殖和转化中的作用

• 批准号: 8244651
• 负责人: Jeffrey A. Frost
• 金额: $ 5.82万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-06-01 至 2013-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8244651
• 关键词: Address; Affect; Automobile Driving; Binding Sites; Breast; Breast Cancer Cell; C-terminal; Cancer Etiology; Cancer cell line; Cell Cycle Progression; Cell Proliferation; Complex; Cytoplasm; DNA; Data; Development; Disease; Enzymes; Epithelial Cell Proliferation; Epithelial Cells; Family; Goals; Guanine Nucleotide Exchange Factors; Human; Human Papillomavirus; Human T-Cell Leukemia Viruses; In Vitro; Intervention; Malignant Neoplasms; Mammary Neoplasms; Mediating; Monomeric GTP-Binding Proteins; Neuroepithelioma; Normal Cell; Normal tissue morphology; Nuclear; Oncogene Proteins; Phosphorylation; Phosphotransferases; Protein Isoforms; Protein Kinase; Proteins; RNA Splicing; Regulation; Research; Role; Site; Tumor Suppressor Proteins; Type I DNA Topoisomerases; Variant; Virus; base; cell motility; cell transformation; genetic regulatory protein; human TOP1 protein; in vitro activity; malignant breast neoplasm; metaplastic cell transformation; overexpression; protein activation; protein complex; rho; tumor

DESCRIPTION (provided by applicant): Rho family small G protein activation has been implicated in the development of cancer and in metastatic progression. Rho protein activation is controlled by a family of enzymes known as guanine nucleotide exchange factors (Rho GEFs), so understanding the mechanisms regulating Rho GEF activity is critical to devising strategies to block Rho protein-mediated transformation. NET1 is a nuclear Rho GEF that is specific for the RhoA subfamily of small G proteins. Overexpression of NET1 results in its mislocalization in the cytoplasm, and stimulates constitutive RhoA activation and cell transformation. We have observed that NET1 isoforms are overexpressed in primary breast tumors, and that overexpression of NET1 proteins in breast cancer cell lines dramatically stimulates their proliferation. We have also observed that NET1 requires a C-terminal PDZ binding site to stimulate cell proliferation and transformation, and that this binding site mediates interaction with a protein complex consisting of the tumor suppressor Dlg1, the related protein Cask, and the DNA modifying enzyme Topo I. Since Dlg1 is a requisite target for oncoproteins from cancer causing viruses such as the human papilloma virus and the human T cell leukemia virus 1, the interaction between NET1 and Dlg1 is likely to be especially important to the mechanism by which NET1 controls cell proliferation. The hypothesis driving the proposed research is that overexpression of NET1 causes its mislocalization in the cytoplasm, and results in the constitutive activation of RhoA and an inhibition of the tumor suppressor function of Dlg1. We will address this hypothesis in the following specific aims. In Aim1 we will characterize the interaction of NET1 isoforms with Dlg1, Cask and Topo I in vitro and in breast epithelial cells. In Aim 2 we will elucidate how the phosphorylation of NET1 isoforms by regulatory kinases controls the interaction of NET1 with Dlg1. In Aim 3 we will examine the effects of NET1 overexpression, complex formation with Dlg1, Cask and Topo I, and NET1 phosphorylation on breast epithelial cell proliferation and transformation. Completion of these aims will delineate a unique and previously unrecognized role for NET1 and Dlg1 in controlling breast epithelial cell transformation, and identify potential new targets for intervention in this disease.

描述(由申请人提供)：Rho家族小G蛋白激活与癌症的发展和转移进展有关。Rho蛋白的激活受一系列被称为鸟嘌呤核苷酸交换因子(Rho GEF)的酶的控制，因此了解调控Rho蛋白活性的机制对于设计阻止Rho蛋白介导的转化的策略至关重要。Net1是一种针对RhoA亚家族小G蛋白的核Rho全环蛋白。Net1的过表达导致其在细胞质中的错误定位，并刺激结构性的RhoA激活和细胞转化。我们观察到Net1亚型在原发乳腺肿瘤中过度表达，并且Net1蛋白在乳腺癌细胞系中的过度表达显著刺激了它们的增殖。我们还观察到，Net1需要一个C-末端的PDZ结合位点来刺激细胞的增殖和转化，并且这个结合位点介导了与由肿瘤抑制因子Dlg1、相关蛋白Cask1和DNA修饰酶Topo I组成的蛋白质复合体的相互作用。由于Dlg1是致癌病毒(如人乳头瘤病毒和人T细胞白血病病毒1)癌蛋白的必需靶点，Net1和Dlg1之间的相互作用可能对Net1控制细胞增殖的机制特别重要。支持这项研究的假设是，Net1的过度表达导致其在细胞质中的错误定位，并导致RhoA的结构性激活和Dlg1的肿瘤抑制功能。我们将在以下具体目标中解决这一假设。在Aim1中，我们将在体外和乳腺上皮细胞中研究Net1亚型与Dlg1、CaskI和Topo I的相互作用。在目标2中，我们将阐明调节激酶对Net1异构体的磷酸化如何控制Net1与Dlg1的相互作用。在目标3中，我们将检测Net1过表达、与Dlg1、CaskI和Topo I形成复合体，以及Net1磷酸化对乳腺上皮细胞增殖和转化的影响。这些目标的完成将勾勒出Net1和Dlg1在控制乳腺上皮细胞转化中的独特和以前未被认识的作用，并确定干预这种疾病的潜在新靶点。

项目成果

Cdk1 phosphorylation negatively regulates the activity of Net1 towards RhoA during mitosis.

• DOI: 10.1016/j.cellsig.2021.109926
• 发表时间: 2021-04
• 期刊: Cellular signalling
• 影响因子: 4.8
• 作者: Ulu A;Oh W;Zuo Y;Frost JA
• 通讯作者: Frost JA

The RhoGEF Net1 is required for normal mammary gland development.

RhoGEF Net1 是正常乳腺发育所必需的。

• DOI: 10.1210/me.2014-1128
• 发表时间: 2014
• 期刊: Molecular endocrinology (Baltimore, Md.)
• 作者: Zuo,Yan;Berdeaux,Rebecca;Frost,JeffreyA
• 通讯作者: Frost,JeffreyA

Coexpression of alpha6beta4 integrin and guanine nucleotide exchange factor Net1 identifies node-positive breast cancer patients at high risk for distant metastasis.

• DOI: 10.1158/1055-9965.epi-08-0842
• 发表时间: 2009-01
• 期刊: Cancer epidemiology, biomarkers & prevention : a publication of the American Association for Cancer Research, cosponsored by the American Society of Preventive Oncology
• 作者: Gilcrease MZ;Kilpatrick SK;Woodward WA;Zhou X;Nicolas MM;Corley LJ;Fuller GN;Tucker SL;Diaz LK;Buchholz TA;Frost JA
• 通讯作者: Frost JA

p21 activated kinase 5 activates Raf-1 and targets it to mitochondria.

• DOI: 10.1002/jcb.21809
• 发表时间: 2008-09-01
• 期刊: JOURNAL OF CELLULAR BIOCHEMISTRY
• 影响因子: 4
• 作者: Wu, Xiaochong;Carr, Heather S.;Dan, Ippeita;Ruvolo, Peter P.;Frost, Jeffrey A.
• 通讯作者: Frost, Jeffrey A.

A bacterial cytotoxin identifies the RhoA exchange factor Net1 as a key effector in the response to DNA damage.

• DOI: 10.1371/journal.pone.0002254
• 发表时间: 2008-05-28
• 期刊: PLOS ONE
• 影响因子: 3.7
• 作者: Guerra, Lina;Carr, Heather S.;Richter-Dahlfors, Agneta;Masucci, Maria G.;Thelestam, Monica;Frost, Jeffrey A.;Frisan, Teresa
• 通讯作者: Frisan, Teresa