Regulation and function of the cerebral cavernous malformation 2 protein Public Deposited

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  • March 21, 2019
  • Crose, Lisa Eileen Stalheim
    • Affiliation: School of Medicine, Department of Pharmacology
  • Cerebral cavernous malformations (CCM) are vascular lesions of the central nervous system characterized as clusters of dilated, thin-walled blood vessels. CCM lesions are fragile and prone to vascular leakiness and rupture, leading to hemorrhages that cause seizure and stroke. Familial CCM has been shown to be genetically linked to three genes: CCM1, CCM2, and CCM3. The proteins encoded by these genes have no apparent catalytic activity, suggesting they are scaffolds to organize and localize functional protein complexes in cells. This scaffolding function has been appreciated for CCM2, which encodes Osmosensing Scaffold for MEKK3 (OSM). CCM2 (OSM) coordinates a signaling complex that consists of Rac1, MEKK3, and MKK3 to activate p38 in response to osmotic stimuli. The studies described here analyze the function of CCM2 in the context of cerebral cavernous malformations. Using proteomic, biochemical, and in vivo models, we characterize CCM2 as a critical regulator of endothelial cell signaling and function. We show that CCM2 binds and localizes the CCM1 protein. The CCM2 phosphotyrosine binding (PTB) domain is necessary for a canonical interaction with NPxY motifs within CCM1. We provide evidence of co-immunoprecipitation and fluorescence resonance energy transfer (FRET) between CCM1 and CCM2, implicating a common genetic and molecular pathway in CCM pathogenesis. We also characterize CCM2 as a Smurf1 binding partner. Through a novel CCM2 PTB domain - Smurf1 HECT domain interaction, CCM2 recruits Smurf1 to specific locations at the plasma membrane where it specifically degrades RhoA. Knockdown of CCM2 in brain endothelial cells leads to increased RhoA protein levels and ROCK signaling. Functionally, this leads to deficiencies in cell migration, tube formation, and maintenance of a permeability barrier. To determine the role of CCM2 in vivo, we used Danio rerio as a model for vertebrate development. Loss of CCM2 expression leads to decreased blood flow due to restrictions and abnormalities of the aortic arch. The findings presented here indicate that CCM2 regulates protein complexes and signaling pathways important in endothelial cell function and provide insight into the molecular mechanisms involved in cerebral cavernous malformation pathogenesis.
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  • Johnson, Gary
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  • University of North Carolina at Chapel Hill
  • Open access

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