Abstract
Proteolytic cleavage and release from the cell surface of membrane-tethered ligands is an important mechanism of regulating intercellular signalling. TACE is a major shedding protease, responsible for the liberation of the inflammatory cytokine TNFα and ligands of the epidermal growth factor receptor. iRhoms, catalytically inactive members of the rhomboid-like superfamily, have been shown to control the ER-to-Golgi transport and maturation of TACE. Here, we reveal that iRhom2 remains associated with TACE throughout the secretory pathway, and is stabilised at the cell surface by this interaction. At the plasma membrane, ERK1/2-mediated phosphorylation and 14-3-3 protein binding of the cytoplasmic amino-terminus of iRhom2 alter its interaction with mature TACE, thereby licensing its proteolytic activity. We show that this molecular mechanism is responsible for triggering inflammatory responses in primary mouse macrophages. Overall, iRhom2 binds to TACE throughout its lifecycle, implying that iRhom2 is a primary regulator of stimulated cytokine and growth factor signalling.
Highlights
Signalling ligands are often synthesised as transmembrane domain (TMD) containing precursors
TACE activity, but not TACE maturation, is regulated by iRhom2 phosphorylation iRhom2 is an essential regulator of TACE maturation and TACE-dependent inflammatory and growth factor signalling (Adrain et al, 2012; McIlwain et al, 2012; Siggs et al, 2012; Christova et al, 2013; Issuree et al, 2013; Li et al, 2015)
Once TACE has been exported from the ER under the influence of iRhom2 (Figure 7a), and its inhibitory pro-domain is removed in the trans-Golgi network, interaction with iRhom2 is essential to stabilise it at the plasma membrane, preventing its lysosomal degradation (Figure 7b)
Summary
Signalling ligands are often synthesised as transmembrane domain (TMD) containing precursors. TACE-induced ligand shedding is rapid: it is activated within ten minutes by physiological and pharmacological stimulation (Le Gall et al, 2010), and this is compatible with the timescale on which iRhoms have been reported to influence TACE substrate selection (Maretzky et al, 2013) This speed of TACE activation is not reconciled with its relatively slow ER-to-Golgi trafficking and maturation (3–6 hr, [Schlondorff et al, 2000]), the process by which iRhoms are known to control TACE. IRhom acts as a signalling hub whereby phosphorylation of its N-terminus and subsequent 14-3-3 protein binding controls stimulated TACE proteolytic activity and ligand shedding We show that this regulation of mature TACE activity occurs in immortalised cell models and regulates inflammatory TNFa release from primary macrophages. We conclude that iRhom binds to TACE and controls its function at multiple stages, indicating that iRhom can be treated as a multifunctional regulatory subunit of TACE
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