1 Distinct roles for Rho vs. Rac/Cdc42 GTPases downstream of Vav2 in regulating mammary epithelial acinar architecture Lei Duan 1, *, Gengsheng Chen 5 , Sumeet Virmani 6 , GuoGuang Ying 7 , , Srikumar M. Raja 1 , Byung Min Chung 1 , Mark A. Rainey 1 , Manjari Dimri 8 , Cesar F. Ortega-Cava 1 , Xiangshan Zhao 3 , Robert J. Clubb 1 , Chun Tu 1 , Alagarsamy L. Reddi 9 , Mayumi Naramura 1,4 , Vimla Band 1,2,4, *, Hamid Band 1,2,3,4, * 1 Eppley Institute for Cancer and Allied Diseases; 2 Department of Genetics, Cell Biology and Anatomy, College of Medicine and 3 Department of Biochemistry and Molecular Biology; and 4 UNMC-Eppley Cancer Center, University of Nebraska Medical Center, 985950 Nebraska Medical Center, Omaha, NE 68198-5950. Current addresses: 5 GC: Coskata Energy, Warrenville, IL; 6 SV: Department of Radiology, Northwestern University Feinberg School of Medicine, Chicago, IL; 7 GY: Tianjin Cancer Hospital Cancer Institute, Tianjin Medical University, West HuanHu Road, HeXi District, Tianjin, P.R.China; 8 MD: NorthShore University HealthSystem Research Institute, Evanston, IL; 9 ALR: Department of Medicine, Northwestern University Feinberg School of Medicine, Chicago, IL Running Head: Vav2 and Rho GTPases control acinar morphogenesis *Address correspondence to : Lei Duan, MD, Vimla Band, Ph.D or Hamid Band, MD, Ph.D, University of Nebraska Medical Center, 986805 Nebraska Medical Center, Omaha, NE 68198- 6805; Phone: 402-559-8568(LD)/8572(HB)/8565(VB); FAX: 402-559-4651; E-mail: lduan@unmc.edu ; vband@unmc.edu ; hband@unmc.edu Non-malignant mammary epithelial cells (MECs) undergo acinar morphogenesis in three-dimensional (3D) Matrigel culture, a trait that is lost upon oncogenic transformation. Rho GTPases are thought to play important roles in regulating epithelial cell-cell junctions, but their contributions to acinar morphogenesis remain unclear. Here we report that the activity of Rho GTPases is downregulated in non-malignant MECs in 3D culture with particular suppression of Rac1 and Cdc42. Inducible expression of a constitutively- active form of Vav2, a Rho GTPase guanine nucleotide exchange factor (GEF) activated by receptor tyrosine kinases, in 3D MEC culture activated Rac1 and Cdc42; Vav2 induction from early stages of culture impaired acinar morphogenesis and induction in pre-formed acini disrupted the pre-established acinar architecture and led to cellular outgrowths . Knockdown studies demonstrated that Rac1 and Cdc42 mediate the constitutively-active Vav2 phenotype, while in contrast, RhoA knockdown intensified the Vav2-induced disruption of acini, leading to more aggressive cell outgrowth and branching morphogenesis. These results indicate that RhoA plays an antagonistic role to Rac1/Cdc42 in the control of mammary epithelial acinar morphogenesis. Differentiated epithelia display a polarized architecture that is essential for their functional role as protective barriers and secretory or absorptive surfaces. The polarized epithelial cells associate with each other through lateral cell-cell junctions, which functionally and biochemically segregate the apical surface from the extracellular matrix (ECM)-contacting basal surface (1,2). The cell-cell junctions and cell- ECM interactions stabilize the epithelial structure and ensure appropriate signaling http://www.jbc.org/cgi/doi/10.1074/jbc.M109.057976 The latest version is at JBC Papers in Press. Published on October 13, 2009 as Manuscript M109.057976 Copyright 2009 by The American Society for Biochemistry and Molecular Biology, Inc. by guest on October 5, 2017 http://www.jbc.org/ Downloaded from