Search

The ubiquitination of IFNAR1 governs the endocytosis of type I interferon (IFN1) receptor and regulates the extent of IFN1 signaling. This ubiquitination is mediated by the βTrcp E3 ubiquitin ligase, which is capable of building both Lys48- and Lys63-linked chains on IFNAR1. Although ubiquitination can occur on many Lys residues within the intracellular domain, a cluster of three lysines (Lys-501, Lys-525 and Lys-526) is required for efficient IFNAR1 endocytosis. Ubiquitination of this cluster acts to unmask an adjacent Tyr-466-based linear endocytic motif and enable its interaction with the AP2 adaptin complex and subsequent IFNAR1 internalization via the clathrin-dependent pathway. However, it remained unclear how both Lys-48- and Lys-63-linked chains are assembled to stimulate IFNAR1 endocytosis. Here we propose a model where the two types of ubiquitin chains stabilize the conformation of the intracellular tail of IFNAR1 to expose Tyr-466 motif that is otherwise masked by TYK2. Whereas the βTrcp facilitates both Lys-48- and Lys-63-polyubiquitination, the specificity of these processes is determined by two alternate E2 ubiquitin conjugating enzymes, Ubc13 and Cdc34. Ubc13 aids βTrcp to build the Lys-63-linked polyubiquitination of Lys-501, whereas the Lys-48-chain-specific Cdc34 is required for modification of that occurs on Lys-525, 526. These data indicate that βTrcp alternates between two diverse E2s to stimulate IFNAR1 complex topology ubiquitination. We propose that this complex ubiquitination relieves the masking of Tyr-466 by relieving a bend in the polypeptide chain conferred by proline residue Pro470 located upstream of ubiquitin acceptor sites. Consistent with this hypothesis, IFNAR1 P470G mutant (in which this turn is removed) undergoes β Trcp-independent constitutive endocytosis despite retaining TYK2 interaction. [ABSTRACT FROM AUTHOR]

Alpha interferon (IFN-α) controls homeostasis of hematopoietic stem cells, regulates antiviral resistance, inhibits angiogenesis, and suppresses tumor growth. This cytokine is often used to treat cancers and chronic viral infections. The extent of cellular responses to IFN-α is limited by the IFN-induced ubiquitination and degradation of the IFN-α/β receptor chain 1 (IFNAR1) chain of the cognate receptor. IFNAR1 ubiquitination is facilitated by the βTrcp E3 ubiquitin ligase that is recruited to IFNAR1 upon its degron phosphorylation, which is induced by the ligand. Here we report identification of protein kinase D2 (PKD2) as a kinase that mediates the ligand-inducible phosphorylation of IFNAR1 degron and enables binding of βTrcp to the receptor. Treatment of cells with IFN-α induces catalytic activity of PKD2 and stimulates its interaction with IFNAR1. Expression and kinase activity of PKD2 are required for the ligand-inducible stimulation of IFNAR1 ubiquitination and endocytosis and for accelerated proteolytic turnover of IFNAR1. Furthermore, inhibition or knockdown of PKD2 robustly augments intracellular signaling induced by IFN-α and increases the efficacy of its antiviral effects. The mechanisms of the ligand-inducible elimination of IFNAR1 are discussed, along with the potential medical significance of this regulation. [ABSTRACT FROM AUTHOR]

Linear endocytic motifs of signaling receptors as well as their ubiquitination determine the rate of ligand-induced endocytosis that mediates down-regulation of these receptors and restricts the magnitude and duration of their respective signal transduction pathways. We previously hypothesized that, in the absence of its cognate ligand, type I interferon (IFN), the IFNalpha receptor chain 1 (IFNAR1) receptor chain is protected from basal endocytosis by a hypothetical masking complex that prevents the Tyr-based endocytic motif within IFNAR1 from interacting with components of the adaptin protein complex 2 (AP2). Here we identify a member of the Janus kinase (Jak) family, Tyk2, as a component of such a masking complex. In the absence of ligand or of receptor chain ubiquitination, binding of Janus kinase Tyk2 within the proximity of the Tyr-based linear motif of IFNAR1 is required to prevent IFNAR1 internalization and to maintain its cell surface expression. Furthermore, interaction experiments revealed that Tyk2 physically shields this Tyr-based motif from the recognition by the AP50 subunit of AP2. These data delineate a long-sought ligand- and ubiquitin-independent mechanism by which Tyk2 contributes to both the regulation of total IFNAR1 levels as well as the regulation of the cell surface density of this receptor chain.

Prolactin (PRL) activates its receptor to initiate signal transduction pathways (including activation of Janus kinases, Jak) but also stimulates downregulation of this receptor to limit the magnitude and duration of signaling. Degradation of the long form of PRL receptor (PRLr) depends on its phosphorylation on Ser349 that is required to facilitate PRLr ubiquitination. Signaling events that mediate PRL-induced degradation of PRLr remain to be elucidated. Here, we investigated the role of Jak2 activity in ligand-triggered increase of PRLr phosphorylation on Ser349, PRLr ubiquitination, endocytosis, and degradation. Using Jak2 reconstitution in Jak2-null cells as well as pharmacologic approaches, we found that treatment with PRL (but not with PRLr antagonist) promotes phosphorylation of PRLr on Ser349 and accelerates endocytosis of PRLr. Furthermore, PRL-stimulated PRLr phosphorylation, endocytosis, and degradation in Jak2-null cells reconstituted with wild type but not with catalytically inactive Jak2. We discuss how Jak2-mediated signaling might be transduced into Ser349 phosphorylation of PRLr as well as its ubiquitination and endocytosis.