Regulated Proteolytic Processing of Tie1 Modulates Ligand Responsiveness of the Receptor-tyrosine Kinase Tie2

Abstract

Regulated ectodomain shedding followed by intramembrane proteolysis has recently been recognized as important in cell signaling and for degradation of several type I transmembrane proteins. The receptor-tyrosine kinase Tie1 is known to undergo ectodomain cleavage generating a membrane-tethered endodomain. Here we show Tie1 is a substrate for regulated intramembrane proteolysis. After Tie1 ectodomain cleavage the newly formed 45-kDa endodomain undergoes additional proteolytic processing mediated by ␥-secretase to generate an amino-terminal-truncated 42-kDa fragment that is subsequently degraded by proteasomal activity. This sequential processing occurs constitutively and is stimulated by phorbol ester and vascular endothelial growth factor. To assess the biological significance of regulated Tie1 processing, we analyzed its effects on angiopoietin signaling. Activation of ectodomain cleavage causes loss of phosphorylated Tie1 holoreceptor and generation of phosphorylated receptor fragments in the presence of cartilage oligomeric protein angiopoietin 1. A key function of ␥-secretase is in preventing accumulation of these phosphorylated fragments. We also find that regulated Tie1 processing modulates ligand responsiveness of the Tie-1-associated receptor Tie2. Activation of Tie1 ectodomain cleavage increases cartilage oligomeric protein angiopoietin 1 activation of Tie2. This correlates with increased ability of Tie2 to bind ligand after shedding of the Tie1 extracellular domain. A similar enhancement of ligand activation of Tie2 is seen when Tie1 expression is suppressed by RNA interference. Together these data indicate that Tie1, via its extracellular domain, limits the ability of ligand to bind and activate Tie2. Furthermore the data suggest that regulated processing of Tie1 may be an important mechanism for controlling signaling by Tie2.Regulated sequential proteolytic processing has recently emerged as an important mechanism in signal transduction and degradation of transmembrane proteins (1, 2). Such processing has been described for a number of transmembrane proteins, including Notch, amyloid precursor protein and the receptortyrosine kinase ErbB-4 and involves an initial metalloproteasemediated ectodomain shedding followed by secondary cleavage of the remaining membrane-associated fragment (3-8). These sequential cleavage events have been designated RIP 4 for regulated intramembrane proteolysis (1, 2).The initiating and key regulatory step in RIP is ectodomain cleavage, and in most cases this is catalyzed by members of a disintegrin and metalloprotease (ADAM) family, although matrix metalloproteases and the aspartyl proteases ␤-site amyloid precursor protein-cleaving enzymes 1 and 2 have also been implicated to a lesser degree (9, 10). For both Notch and ErbB-4 ectodomain shedding requires ADAM17, also known as tumor necrosis factor-␣-converting enzyme (TACE) (4, 11). This protease has also been implicated in ␤-amyloid precursor protein cleavage (6). After loss of ectodomain, the remaining transmembrane and ...

Receptor Tyrosine Kinases in the Nucleus

“…Ectodomain cleavage of Tie 1 is stimulated by a variety of agents (TPA, VEGF, TNFa, sheer stress) and increases Ang 1 activation of Tie 2, apparently by allowing greater access of the ligand to its Tie 2 binding site. Following ectodomain release, the Tie 1 cell-associated 45-kDa cleavage fragment is processed by g-secretase to produce a 42-kDa cytoplasmic ICD fragment (Marron et al 2007). In this receptor system, the ectodomain secretase action is physiologically important; however, the significance of g-secretase activity may be simply to remove the highly tyrosine phosphorylated 45 kDa fragment.…”

Proteolytic cleavages give receptor tyrosine kinases the gift of ubiquity

“…Downregulation of RTK through PS-RIP also affects the associated signalling pathways. Indeed, as recently shown in the case of the RTK, Tie1, its proteolyses by PS-RIP leads to an amplified ligand activation of the associated receptor, Tie2 (Marron et al, 2007;McCarthy et al, 1999;Yabkowitz et al, 1999). The authors propose that release of the extracellular domain of Tie1 permits ligand access to Tie2, presumably by preventing spatial hindrance.…”

“…However, for numerous RTK, truncation of the extracellular domain leads to the generation of active proteins often showing transforming properties. For IGFR, ErbB4 and Tie1, ectodomain shedding leads to the generation of constitutively phosphorylated membrane-tethered fragments, but their accumulation is prevented by g-secretase cleavage and subsequent degradation of the fragment Marron et al, 2007;McElroy et al, 2007). Thus, g-secretase cleavage might be considered as a protective cleavage preventing the generation of transforming fragments.…”

“…As evidenced, for instance, for CSF-R, IGF-IR, Tie1, EphB2 and Met the ICD is rapidly degraded through a proteasome-dependent mechanism (Wilhelmsen and van der Geer, 2004;Litterst et al, 2007;Marron et al, 2007;McElroy et al, 2007;Foveau et al, 2009) (Figure 2). Moreover, PS-RIP shares common features with the liganddependent degradation described for RTK, including endocytosis and ubiquitination.…”

Temporal expression in rats of receptor tyrosine kinase Tie2 during early wound healing after tooth extraction

“…Temporal expression in rats of receptor tyrosine kinase Tie2 during early wound healing after tooth extraction Tie2 signaling (16,17), with Tie1-Tie2 complex formation (18), causing detachment of endothelial cells from pericytes. This detachment in turn enables endothelial cells to proliferate in the presence of vascular endothelial growth factor (14,19,20).…”