Research in the Glading lab focuses on the role of cell-cell contact in regulating cellular function in both the normal and disease state. Specifically, we are interested in studying the molecular mechanisms that regulate the integrity and growth of blood vessels. A second major focus of the lab is the regulation of cell-cell contacts between epithelial cells, and how those contacts are disrupted during tumor growth and metastasis. We use cell culture and animal models to investigate specific signaling pathways involved in these processes.

Our research is currently focused on a new family of proteins that we have identified as regulators of cell-cell contact. The CCM proteins are scaffolding proteins that organize an intracellular complex at the cell-cell junction. Loss of any of the CCM proteins (KRIT1 (also called CCM1), CCM2 (malcavernin/OSM), and CCM3 (PDCD10)) leads to the development of Cerebral Cavernous Malformation (CCM), a type of vascular malformation found in 0.1-0.5% of the population. This disease is characterized by the development of focal areas of abnormal capillary growth within the brain and other tissues. CCM lesions consist of beds of dilated, leaky blood vessels, with defective endothelial cell-cell junctions. Symptoms include seizure, cerebral hemorrhage and focal neurological deficits, likely due to blood leakage from the lesion into the surrounding tissue.

Of the CCM proteins, we identified KRIT1 as a component of cell-cell junctions, where it interacts with structural and signaling proteins to regulate the stability of the junction. KRIT1 was first identified as a binding partner of the GTPase Rap1a, and we have shown that KRIT1 is the Rap1 effector responsible for the ability of active Rap1 to stabilize endothelial cell-cell adhesion. Loss of KRIT1 directly contributes to permeability and vascular leak by inhibiting the association of beta- and p120-catenin with VE-cadherin, and by altering the activity of Rho family GTPases. Finally, loss of KRIT1, by de-stabilizing cell-cell adhesion, promotes the localization of beta-catenin to the nucleus, where it modulates gene expression. However, many questions remain unanswered including; 1) what are the mechanisms that regulate KRIT1 signaling in normal cells? and 2) how does KRIT1 signaling contribute to changes in cellular behavior?

Although CCM is type of vascular malformation, KRIT1 is a ubiquitous protein, and our evidence indicates that it regulates cell-cell contacts in many types of cells. As the dynamic regulation of cell-cell contact, including beta-catenin signaling, is important for tissue maintenance and growth, the outcome of this research has the potential to impact a wide range of scientific fields. For example, our data indicate that KRIT1 functions as a tumor suppressor in epithelial cells, a novel function for this family of proteins. Our exciting studies continue to explore the molecular mechanism(s) behind this newly described function, as well as probe the contribution of KRIT1 signaling to the pathogenesis of other diseases reliant on vascular leak and beta-catenin signaling, such as arthritis.

• Corr M, Lerman I, Keubel JM, Ronacher L, Misra R, Lund F, Sarelius IH, Glading AJ. Decreased krev interaction-trapped 1 expression leads to increased vascular permeability and modifies inflammatory responses in vivo. Arterioscler Thromb Vasc Biol. 2012 Nov; 32(11):2702-10.
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• Glading AJ, Ginsberg MH. Rap1 and its effector KRIT1/CCM1 regulate beta-catenin signaling. Dis Model Mech. 2010 Jan-Feb; 3(1-2):73-83.
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• Glading A, Han J, Stockton RA, Ginsberg MH. KRIT-1/CCM1 is a Rap1 effector that regulates endothelial cell cell junctions. J Cell Biol. 2007 Oct 22; 179(2):247-54.
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• Glading A, Koziol JA, Krueger J, Ginsberg MH. PEA-15 inhibits tumor cell invasion by binding to extracellular signal-regulated kinase 1/2. Cancer Res. 2007 Feb 15; 67(4):1536-44.
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• Krueger J, Chou FL, Glading A, Schaefer E, Ginsberg MH. Phosphorylation of phosphoprotein enriched in astrocytes (PEA-15) regulates extracellular signal-regulated kinase-dependent transcription and cell proliferation. Mol Biol Cell. 2005 Aug; 16(8):3552-61.
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• Satish L, Blair HC, Glading A, Wells A. Interferon-inducible protein 9 (CXCL11)-induced cell motility in keratinocytes requires calcium flux-dependent activation of mu-calpain. Mol Cell Biol. 2005 Mar; 25(5):1922-41.
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• Glading A, Bodnar RJ, Reynolds IJ, Shiraha H, Satish L, Potter DA, Blair HC, Wells A. Epidermal growth factor activates m-calpain (calpain II), at least in part, by extracellular signal-regulated kinase-mediated phosphorylation. Mol Cell Biol. 2004 Mar; 24(6):2499-512.
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• Chou FL, Hill JM, Hsieh JC, Pouyssegur J, Brunet A, Glading A, Uberall F, Ramos JW, Werner MH, Ginsberg MH. PEA-15 binding to ERK1/2 MAPKs is required for its modulation of integrin activation. J Biol Chem. 2003 Dec 26; 278(52):52587-97.
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• Shiraha H, Glading A, Chou J, Jia Z, Wells A. Activation of m-calpain (calpain II) by epidermal growth factor is limited by protein kinase A phosphorylation of m-calpain. Mol Cell Biol. 2002 Apr; 22(8):2716-27.
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• Glading A, Lauffenburger DA, Wells A. Cutting to the chase: calpain proteases in cell motility. Trends Cell Biol. 2002 Jan; 12(1):46-54.
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• Glading A, Uberall F, Keyse SM, Lauffenburger DA, Wells A. Membrane proximal ERK signaling is required for M-calpain activation downstream of epidermal growth factor receptor signaling. J Biol Chem. 2001 Jun 29; 276(26):23341-8.
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• Glading A, Chang P, Lauffenburger DA, Wells A. Epidermal growth factor receptor activation of calpain is required for fibroblast motility and occurs via an ERK/MAP kinase signaling pathway. J Biol Chem. 2000 Jan 28; 275(4):2390-8.
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• Shiraha H, Glading A, Gupta K, Wells A. IP-10 inhibits epidermal growth factor-induced motility by decreasing epidermal growth factor receptor-mediated calpain activity. J Cell Biol. 1999 Jul 12; 146(1):243-54.
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• Wells A, Gupta K, Chang P, Swindle S, Glading A, Shiraha H. Epidermal growth factor receptor-mediated motility in fibroblasts. Microsc Res Tech. 1998 Dec 1; 43(5):395-411.
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