Angiogenesis is essential for normal physiological processes such as organ development and wound healing. It is also critical to pathological processes such as tumor growth, atherosclerosis, rheumatoid arthritis, and diabetic retinopathy. Therefore, blocking angiogenesis could be a powerful therapeutic intervention for treating diseases requiring formation of new blood vessels. However, the underlying molecular mechanism of intracellular signal transduction in this process remains largely unexplored. We recently generated Mekk3-knockout mice through homologous recombination and demonstrated that MEKK3, a Ser/Thr protein kinase belonging to the mitogen-activated protein kinase (MAPK) kinase gene family, is essential for angiogenesis. The long-term goals of this study are to elucidate the molecular mechanisms of angiogenesis regulated by MEKK3 signaling.
Our SPECIFIC AIM 1 is to determine the function of MEKK3 in endothelial cells (ECs) during embryonic development by examining the morphology, proliferation and apoptosis of ECs in the E8.5 to E10 wild-type, Mekk3+/- and Mekk3-/- fetuses. We will also isolate embryonic ECs from the E9-E9.5 fetuses and establish EC lines by using polyoma middle T antigen to transform the primary cultures.
In SPECIFIC AIM 2, we will determine whether MEKK3 is a specific upstream activator of JNK1/2, ERK1/2, p38 and ERK5 MAPKs in ECs by using in vitro kinase assays. We will determine whether the induction of these MAPKs is defective in Mekk3-/- embryos, purified ECs and EC lines. In addition, we will determine whether MEKK3 is specifically activated by angiogenic stimulation by using in vitro kinase, in-gel kinase and MEKK3 phosphorylation assays. Furthermore, we will isolate and clone MEKK3-associated proteins by coprecipitation, chromatography, and yeast two-hybrid screening.
In Specific Aim 4, we plan to generate Mekk3 floxed ES cells and mice, and use these mice to create EC-specific Mekk3 knock out (Mekk3 (EC-null)) mice and Mekk3-deficient ECs with Cre recombinase. Function of MEKK3-signaling will be studied specifically in ECs using these mice. In addition to revealing the MEKK3-signaling in angiogenesis, the outcome from this study will provide conceptual and material resources for studying MEKK3 and its homologues in many other physiological and pathological processes. Most importantly, this study may discover novel molecular targets for therapeutic intervention for treating diseases requiring formation of new blood vessels like cancer.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
1R01HL070225-01A1
Application #
6580577
Study Section
Experimental Cardiovascular Sciences Study Section (ECS)
Program Officer
Goldman, Stephen
Project Start
2003-04-01
Project End
2007-03-31
Budget Start
2003-04-01
Budget End
2004-03-31
Support Year
1
Fiscal Year
2003
Total Cost
$325,470
Indirect Cost
Name
University of Texas MD Anderson Cancer Center
Department
Microbiology/Immun/Virology
Type
Other Domestic Higher Education
DUNS #
800772139
City
Houston
State
TX
Country
United States
Zip Code
77030
Ajibade, Adebusola Alagbala; Wang, Qinfu; Cui, Jun et al. (2012) TAK1 negatively regulates NF-?B and p38 MAP kinase activation in Gr-1+CD11b+ neutrophils. Immunity 36:43-54
Chang, Xing; Lazorchak, Adam S; Liu, Dou et al. (2012) Sin1 regulates Treg-cell development but is not required for T-cell growth and proliferation. Eur J Immunol 42:1639-47
Wang, Xiaofang; Zhang, Fan; Chen, Fanping et al. (2011) MEKK3 regulates IFN-gamma production in T cells through the Rac1/2-dependent MAPK cascades. J Immunol 186:5791-800
Chang, Xing; Liu, Fang; Wang, Xiaofang et al. (2011) The kinases MEKK2 and MEKK3 regulate transforming growth factor-?-mediated helper T cell differentiation. Immunity 34:201-12
Yu, Ling; Su, Bing; Hollomon, Mario et al. (2010) Vasculogenesis driven by bone marrow-derived cells is essential for growth of Ewing's sarcomas. Cancer Res 70:1334-43
Lazorchak, Adam S; Liu, Dou; Facchinetti, Valeria et al. (2010) Sin1-mTORC2 suppresses rag and il7r gene expression through Akt2 in B cells. Mol Cell 39:433-43
Oh, Won Jun; Wu, Chang-chih; Kim, Sung Jin et al. (2010) mTORC2 can associate with ribosomes to promote cotranslational phosphorylation and stability of nascent Akt polypeptide. EMBO J 29:3939-51
Facchinetti, Valeria; Ouyang, Weiming; Wei, Hua et al. (2008) The mammalian target of rapamycin complex 2 controls folding and stability of Akt and protein kinase C. EMBO J 27:1932-43
Kim, Kihwan; Duramad, Omar; Qin, Xiao-Feng et al. (2007) MEKK3 is essential for lipopolysaccharide-induced interleukin-6 and granulocyte-macrophage colony-stimulating factor production in macrophages. Immunology 120:242-50
Deng, Yong; Yang, Jianhua; McCarty, Marya et al. (2007) MEKK3 is required for endothelium function but is not essential for tumor growth and angiogenesis. Am J Physiol Cell Physiol 293:C1404-11

Showing the most recent 10 out of 16 publications