The angiopoietins comprise a small class of secreted glycoproteins that play crucial roles in the maturation and maintenance of the mammalian vascular and lymphatic systems. They exert their effects through a member of the tyrosine kinase receptor family, Tie2. Angiopoietin/Tie2 signaling is unique among tyrosine kinase receptor-ligand systems in that distinct angiopoietin ligands, although highly homologous, can function as agonists or antagonists in a context-dependent manner. In an effort to understand this molecular dichotomy, we have crystallized and determined the 2.4 A crystal structure of the Angiopoietin-2 (Ang2) receptor binding region. The structure reveals a fibrinogen fold with a unique C-terminal P domain. Conservation analysis and structure-based mutagenesis identify a groove on the Ang2 molecular surface that mediates receptor recognition.
"Ang1 and Ang2 share approximately 60% homology in their amino acid sequence, as well as a common protein structure consisting of an amino terminal that modulates super-clustering, a coiled-coil domain for the formation of homodimers and a fibrinogen-like domain with ligand activity [16, 40, 132]. Ang1 and Ang2 bind to the same domain on Tie2 with similar affinities [15, 16, 132] and conformation . However, they have opposite effects on receptor phosphorylation and activation. "
[Show abstract][Hide abstract] ABSTRACT: Spinal cord injuries (SCI) can result in devastating paralysis, for which there is currently no robustly efficacious neuroprotective/neuroregenerative treatment. When the spinal cord is subjected to a traumatic injury, the local vasculature is disrupted and the blood-spinal cord barrier is compromised. Subsequent inflammation and ischemia may then contribute to further secondary damage, exacerbating neurological deficits. Therefore, understanding the vascular response to SCI and the molecular elements that regulate angiogenesis has considerable relevance from a therapeutic standpoint. In this paper, we review the nature of vascular damage after traumatic SCI and what is known about the role that angiogenic proteins-angiopoietin 1 (Ang1), angiopoietin 2 (Ang2) and angiogenin-may play in the subsequent response. To this, we add recent work that we have conducted in measuring these proteins in the cerebrospinal fluid (CSF) and serum after acute SCI in human patients. Intrathecal catheters were installed in 15 acute SCI patients within 48 h of injury. CSF and serum samples were collected over the following 3-5 days and analysed for Ang1, Ang2 and angiogenin protein levels using a standard ELISA technique. This represents the first description of the endogenous expression of these proteins in an acute human SCI setting.
Translational Stroke Research 12/2011; 2(4):474-491. DOI:10.1007/s12975-011-0109-x · 2.44 Impact Factor
"Interestingly, our crystal structure of Tie2 bound to Ang2(Barton et al., 2006) and to Ang1 (unpublished data) show that all angiopoietins bind Tie2 in a similar conformation, excluding the possibility of differential Tie2 activation resulting from alternate Tie2/angiopoietin structural arrangements. However, our structural analysis does suggest that the different angiopoietin ligands could present distinct molecular surfaces outside of the receptor-binding interface that could influence the Tie1-Tie2 receptor complexes(Barton et al., 2005). Based upon our findings we hypothesized that the observed direct Tie2/Tie1 interactions are inhibitory and the ability, or inability, of individual angiopoietins to effectively destabilize the Tie1/Tie2 complexes at the cell surface would define their respective agonistic or antagonistic roles. "
[Show abstract][Hide abstract] ABSTRACT: The Tie family of endothelial-specific receptor tyrosine kinases is essential for cell proliferation, migration, and survival during angiogenesis. Despite considerable similarity, experiments with Tie1- or Tie2-deficient mice highlight distinct functions for these receptors in vivo. The Tie2 receptor is further unique with respect to its structurally homologous ligands. Angiopoietin-2 and -3 can function as agonists or antagonists; angiopoietin-1 and -4 are constitutive agonists. To address the role of Tie1 in angiopoietin-mediated Tie2 signaling and determine the basis for the behavior of the individual angiopoietins, we used an in vivo FRET-based proximity assay to monitor Tie1 and -2 localization and association. We provide evidence for Tie1-Tie2 complex formation on the cell surface and identify molecular surface areas essential for receptor-receptor recognition. We further demonstrate that the Tie1-Tie2 interactions are dynamic, inhibitory, and differentially modulated by angiopoietin-1 and -2. Based on the available data, we propose a unified model for angiopoietin-induced Tie2 signaling.
"The P-domain mediates interaction of FReD containing proteins such as fibrinogen and tachylectin 5A with their ligands and it is this region of the Ang FReD that interacts with Tie2 . The structure of Ang2 bound to Tie2 ectodomain has been solved and shows the Ang2 P-domain interacting with the second Ig domain of the receptor which is within a globular fold containing three Ig and three EGF-like domains at the amino-terminal head of the receptor [9–11]. Comparison of un-liganded and Ang2-bound Tie2 ectodomain suggests no significant conformational change in Tie2 on ligand binding . "
[Show abstract][Hide abstract] ABSTRACT: Angiopoietin-1 (Ang1) and Ang2 are ligands for the receptor tyrosine kinase Tie2. Structural data suggest that the two ligands bind Tie2 similarly. However, in endothelial cells Ang1 activates Tie2 whereas Ang2 can act as an apparent antagonist. In addition, each ligand exhibits distinct kinetics of release following binding. These observations suggest that additional factors influence function and binding of angiopoietins with receptors in the cellular context. Previous work has shown that Ang1 binding and activation of Tie2 are inhibited by Tie1, a related receptor that complexes with Tie2 in cells. In this study we have investigated binding of Ang1 and Ang2 to Tie2 in endothelial cells. In contrast to Ang1, binding of Ang2 to Tie2 was found to be not affected by Tie1. Neither PMA-induced Tie1 ectodomain cleavage nor suppression of Tie1 expression by siRNA affected the ability of Ang2 to bind Tie2. Analysis of the level of Tie1 co-immunoprecipitating with angiopoietin-bound Tie2 demonstrated that Ang2 can bind Tie2 in Tie2:Tie1 complexes whereas Ang1 preferentially binds non-complexed Tie2. Stimulation of Tie1 ectodomain cleavage did not increase the agonist activity of Ang2 for Tie2. Similarly, the Tie2-agonist activity of Ang2 was not affected by siRNA suppression of Tie1 expression. Consistent with previous reports, loss of Tie1 ectodomain enhanced the agonist activity of Ang1 for Tie2. Importantly, Ang2 was still able to antagonize the elevated Ang1-activation of Tie2 that occurs on Tie1 ectodomain loss. Together these data demonstrate that Ang1 and Ang2 bind differently to Tie2 at the cell surface and this is controlled by Tie1. This differential regulation of angiopoietin binding allows control of Tie2 activation response to Ang1 without affecting Ang2 agonist activity and maintains the ability of Ang2 to antagonize even the enhanced Ang1 activation of Tie2 that occurs on loss of Tie1 ectodomain. This provides a mechanism by which signalling through Tie2 can be modified by stimuli in the cellular microenvironment.
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