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Mechanism for activation of the EGF receptor catalytic domain by the juxtamembrane segment.

ABSTRACT: Signaling by the epidermal growth factor receptor requires an allosteric interaction between the kinase domains of two receptors, whereby one activates the other. We show that the intracellular juxtamembrane segment of the receptor, known to potentiate kinase activity, is able to dimerize the kinase domains. The C-terminal half of the juxtamembrane segment latches the activated kinase domain to the activator, and the N-terminal half of this segment further potentiates dimerization, most likely by forming an antiparallel helical dimer that engages the transmembrane helices of the activated receptor. Our data are consistent with a mechanism in which the extracellular domains block the intrinsic ability of the transmembrane and cytoplasmic domains to dimerize and activate, with ligand binding releasing this block. The formation of the activating juxtamembrane latch is prevented by the C-terminal tails in a structure of an inactive kinase domain dimer, suggesting how alternative dimers can prevent ligand-independent activation.

SUBMITTER: Jura N

PROVIDER: S-EPMC2814540 | biostudies-literature |

REPOSITORIES: biostudies-literature

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The juxtamembrane region of the EGF receptor functions as an activation domain.

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| S-EPMC4036362 | biostudies-literature

The novel SAM domain protein Aveugle is required for Raf activation in the Drosophila EGF receptor signaling pathway.

Project description:Activation of the Raf kinase by GTP-bound Ras is a poorly understood step in receptor tyrosine kinase signaling pathways. One such pathway, the epidermal growth factor receptor (EGFR) pathway, is critical for cell differentiation, survival, and cell cycle regulation in many systems, including the Drosophila eye. We have identified a mutation in a novel gene, aveugle, based on its requirement for normal photoreceptor differentiation. The phenotypes of aveugle mutant cells in the eye and wing imaginal discs resemble those caused by reduction of EGFR pathway function. We show that aveugle is required between ras and raf for EGFR signaling in the eye and for mitogen-activated protein kinase phosphorylation in cell culture. aveugle encodes a small protein with a sterile alpha motif (SAM) domain that can physically interact with the scaffold protein connector enhancer of Ksr (Cnk). We propose that Aveugle acts together with Cnk to promote Raf activation, perhaps by recruiting an activating kinase.

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Project description:BACKGROUND: Several receptor tyrosine kinases (RTKs) such as EGFR, FGFR, TRK, and VEGFR are capable of localizing in the cell nucleus in addition to their usual plasma membrane localization. Recent reports also demonstrate that nuclear-localized RTKs have important cellular functions such as transcriptional activation. On the basis of preliminary bioinformatic analysis, additional RTKs, including receptor tyrosine kinase-like orphan receptor 1 (Ror1) were predicted to have the potential for nuclear subcellular localization. Ror1 is a receptor protein tyrosine kinase that modulates neurite growth in the central nervous system. Because the nuclear localization capability of the Ror1 cytoplasmic domain has not been reported, we examined the cellular expression distribution of this region. RESULTS: The Ror1 cytoplasmic region was amplified and cloned into reporter constructs with fluorescent tags. Following transfection, the nuclear distribution patterns of transiently expressed fusion proteins were observed. Serial deletion constructs were then used to map the juxtamembrane domain of Ror1 (aa_471-513) for this nuclear translocation activity. Further site-directed mutagenesis suggested that a KxxK-16 aa-KxxK sequence at residues 486-509 is responsible for the nuclear translocation interaction. Subsequent immunofluorescence analysis by cotransfection of Ran and Ror1 implied that the nuclear translocation event of Ror1 might be mediated through the Ran pathway. CONCLUSIONS: We have predicted several RTKs that contain the nuclear localization signals. This is the first report to suggest that the juxtamembrane domain of the Ror1 cytoplasmic region mediates the translocation event. Ran GTPase is also implicated in this event. Our study might be beneficial in future research to understand the Ror1 biological signaling pathway.

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Project description:The epidermal growth factor receptor (EGFR) has several functions in mammalian development and disease, particularly cancer. Most EGF ligands are synthesized as membrane-tethered precursors, and their proteolytic release activates signalling. In Drosophila, rhomboid intramembrane proteases catalyse the release of EGF-family ligands; however, in mammals this seems to be primarily achieved by ADAM-family metalloproteases. We report here that EGF is an efficient substrate of the mammalian rhomboid RHBDL2. RHBDL2 cleaves EGF just outside its transmembrane domain, thereby facilitating its secretion and triggering activation of the EGFR. We have identified endogenous RHBDL2 activity in several tumour cell lines.

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Project description:The human EGF receptor (HER) 2 receptor tyrosine kinase is a survival factor for human cardiomyocytes, and its inhibition may explain the increased incidence of cardiomyopathy associated with the anti-HER2 monoclonal antibody trastuzumab (Genentech, South San Francisco, CA), particularly in patients with prior exposure to cardiotoxic chemotherapies e.g., anthracyclines. Here, we show that GW2974 (HER2/EGF receptor tyrosine kinase inhibitor), but not trastuzumab, activates AMP-activated protein kinase (AMPK), initiating a metabolic stress response in human cardiomyocytes that protects against TNFalpha-induced cell death. GW2974 stimulates calcium dependent fatty acid oxidation in vitro and in the myocardium of GW2974-treated rodents. Calcium chelation or siRNA-targeted AMPK knockdown blocks GW2974 induced fatty acid oxidation. In addition, inhibition of AMPK by a specific inhibitor resulted in increased killing of cardiomyocytes. Elucidating the effects of HER2-targeted therapies on AMPK may predict for risk of cardiomyopathy and provide a novel HER2-targeted strategy designed to protect myocardium from the pro-apoptotic effects of pro-inflammatory cytokines released in response to cardiac injury by chemotherapy or acute ischemia.

| S-EPMC1965560 | biostudies-literature

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