Альтернативные убиквитин-конъюгирующие ферменты Е2 регулируют эндоцитоз рецептора интерферона-1

Ubiquitination of signaling receptors triggers their endocytosis to restrict the extent of cell signaling. Type 1 interferon (IFN1) eliminates its receptor from cell surface via stimulating the ubiquitination of its IFNAR1 chain. While it was suggested that this ubiquitination aids IFNAR1 internalization via relieving a steric hindrance of a linear motif within IFNAR1 from the endocytic machinery, the mechanisms involved remain poorly understood. Here we describe a specific role for two disparate ubiquitin acceptor sites within this receptor. These sites, Lys501 and Lys525 / 526, exhibit a preference for polyubiquitination via either Lys63- or Lys48‑linked chains (K63‑Ub and K48‑Ub, respectively). Whereas the SCFβTrcp E3 ubiquitin ligase controls either type of ubiquitination-dependent IFNAR1 endocytosis, the specificity of these processes is determined by two different E2 ubiquitin conjugating enzymes, Ubc13 and Cdc34. These enzymes can be directly used by SCFβTrcp E3 ubiquitin ligase to generate either K63‑Ub or K48‑Ub in vitro. Ubc13 is involved in IFNAR1 endocytosis driven by the K63‑Ub modification of Lys501, whereas the K48‑Ub-specific Cdc34 affects receptor endocytosis via ubiquitin conjugation that occurs onLys525 / 526. Both types of linkages combine to maximize IFNAR1 endocytosis otherwise suppressed by unfavorable conformation dependent on the presence of a conserved Pro470 within the intracellular domain of IFNAR1. We propose a model where alternate utilization of both E2s to assemble diverse polyubiquitin linkages cooperates to achieve IFNAR1 intracellular domain conformations and spatial arrangements that favor a maximal rate of receptor endocytosis.Убиквитинирование сигнальных рецепторов, вызывающее их эндоцитоз, направлено на подавление передачи сигнала. Деградация рецептора интерферона 1‑го типа (IFN1) на поверхности клетки осуществляется путем убиквитинирования комплекса лиганда с рецептором (IFNAR1). Принято считать, что убиквитинирование способствует взаимодействию между линейным мотивом комплекса IFNAR1 и соответствующими структурами системы эндоцитоза, однако механизм этого процесса остается неясным. В данной работе изучена роль двух различных акцепторных сайтов убиквитина на этом рецепторе. Предпочтительное полиубиквитинирование сайтов Lys501 и Lys525 / 526 обеспечивается посредством Lys63- или Lys48‑связанных цепей (K63‑Ub и K48‑Ub соответственно). Несмотря на то, что убиквитинлигаза SCFβTrcp E3 контролирует оба типа убиквитин-зависимого эндоцитоза IFNAR1, специфика этих процессов определяется двумя различными убиквитин-конъюгирующими ферментами E2 – Ubc13 и Cdc34. Эти ферменты могут непосредственно использоваться убиквитинлигазой SCFβTrcp E3 для создания K63‑Ub или K48‑Ub in vitro. Ubc13 принимает участие в эндоцитозе IFNAR1 путем модификации Lys501 с помощью K63‑Ub, в то время как K48‑Ub-специфичный Cdc34 изменяет эндоцитоз посредством конъюгации с убиквитином, которая происходит на Lys525 / 526. Совместный эффект обоих воздействий максимально стимулирует эндоцитоз IFNAR1, который обычно ингибирован конформационным несоответствием, связанным с наличием консервативного Pro470 во внутриклеточном домене IFNAR1. Мы предлагаем модель, в которой эффекты обоих ферментов E2 объединяют отдельные составляющие системы полиубиквитинирования, обеспечивая им взаимодействие с внутриклеточным доменом IFNAR1 при оптимальном пространственном расположении, что дает наибольшую скорость эндоцитоза рецептора

Endocytosis of the IFNAR1 chain of Type 1 interferon receptor is regulated by diverse E2 ubiquitin conjugation enzymes

Ubiquitination of signaling receptors triggers their endocytosis to restrict the extent of cell signaling. Type 1 interferon (IFN1) eliminates its receptor from cell surface via stimulating the ubiquitination of its IFNAR1 chain. While it was suggested that this ubiquitination aids IFNAR1internalization via relieving a steric hindrance of a linear motif within IFNAR1 from the endocytic machinery, the mechanisms involved remain poorly understood. Here we describe a specific role for two disparate ubiquitin acceptor sites within this receptor. These sites, Lys501 and Lys525 / 526, exhibit a preference for polyubiquitination via either Lys63- or Lys48‑linked chains (K63‑Ub and K48‑Ub, respectively). Whereas the SCFβTrcp E3 ubiquitin ligase controls either type of ubiquitination-dependent IFNAR1 endocytosis, the specificity of these processes is determined by two different E2 ubiquitin conjugating enzymes, Ubc13 and Cdc34. These enzymes can be directly used by SCFβTrcp E3 ubiquitin ligase to generate either K63‑Ub or K48‑Ub in vitro. Ubc13 is involved in IFNAR1 endocytosis driven by the K63‑Ub modification of Lys501, whereas the K48‑Ub-specific Cdc34 affects receptor endocytosis via ubiquitin conjugation that occurs on Lys525 / 526. Both types of linkages combine to maximize IFNAR1 endocytosis otherwise suppressed by unfavorable conformation dependent on the presence of a conserved Pro470 within the intracellular domain of IFNAR1. We propose a model where alternate utilization of both E2s to assemble diverse polyubiquitin linkages cooperates to achieve IFNAR1 intracellular domain conformations and spatial arrangements that favor a maximal rate of receptor endocytosis

ATF4 couples MYC-dependent translational activity to bioenergetic demands during tumour progression.

The c-Myc oncogene drives malignant progression and induces robust anabolic and proliferative programmes leading to intrinsic stress. The mechanisms enabling adaptation to MYC-induced stress are not fully understood. Here we reveal an essential role for activating transcription factor 4 (ATF4) in survival following MYC activation. MYC upregulates ATF4 by activating general control nonderepressible 2 (GCN2) kinase through uncharged transfer RNAs. Subsequently, ATF4 co-occupies promoter regions of over 30 MYC-target genes, primarily those regulating amino acid and protein synthesis, including eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1), a negative regulator of translation. 4E-BP1 relieves MYC-induced proteotoxic stress and is essential to balance protein synthesis. 4E-BP1 activity is negatively regulated by mammalian target of rapamycin complex 1 (mTORC1)-dependent phosphorylation and inhibition of mTORC1 signalling rescues ATF4-deficient cells from MYC-induced endoplasmic reticulum stress. Acute deletion of ATF4 significantly delays MYC-driven tumour progression and increases survival in mouse models. Our results establish ATF4 as a cellular rheostat of MYC activity, which ensures that enhanced translation rates are compatible with survival and tumour progression

Site-specific ubiquitination exposes a linear motif to promote interferon-α receptor endocytosis

Ligand-induced endocytosis and lysosomal degradation of cognate receptors regulate the extent of cell signaling. Along with linear endocytic motifs that recruit the adaptin protein complex 2 (AP2)–clathrin molecules, monoubiquitination of receptors has emerged as a major endocytic signal. By investigating ubiquitin-dependent lysosomal degradation of the interferon (IFN)-α/β receptor 1 (IFNAR1) subunit of the type I IFN receptor, we reveal that IFNAR1 is polyubiquitinated via both Lys48- and Lys63-linked chains. The SCFβTrcp (Skp1–Cullin1–F-box complex) E3 ubiquitin ligase that mediates IFNAR1 ubiquitination and degradation in cells can conjugate both types of chains in vitro. Although either polyubiquitin linkage suffices for postinternalization sorting, both types of chains are necessary but not sufficient for robust IFNAR1 turnover and internalization. These processes also depend on the proximity of ubiquitin-acceptor lysines to a linear endocytic motif and on its integrity. Furthermore, ubiquitination of IFNAR1 promotes its interaction with the AP2 adaptin complex that is required for the robust internalization of IFNAR1, implicating cooperation between site-specific ubiquitination and the linear endocytic motif in regulating this process

Stressor- and Corticotropin releasing Factor-induced Reinstatement and Active Stress-related Behavioral Responses are Augmented Following Long-access Cocaine Self-administration by Rats

Rationale Stressful events during periods of drug abstinence likely contribute to relapse in cocaine-dependent individuals. Excessive cocaine use may increase susceptibility to stressor-induced relapse through alterations in brain corticotropin-releasing factor (CRF) responsiveness. Objectives This study examined stressor- and CRF-induced cocaine seeking and other stress-related behaviors in rats with different histories of cocaine self-administration (SA). Materials and methods Rats self-administered cocaine under short-access (ShA; 2 h daily) or long-access (LgA; 6 h daily) conditions for 14 days or were provided access to saline and were tested for reinstatement by a stressor (electric footshock), cocaine or an icv injection of CRF and for behavioral responsiveness on the elevated plus maze, in a novel environment and in the light–dark box after a 14- to 17-day extinction/withdrawal period. Results LgA rats showed escalating patterns of cocaine SA and were more susceptible to reinstatement by cocaine, EFS, or icv CRF than ShA rats. Overall, cocaine SA increased activity in the center field of a novel environment, on the open arms of the elevated plus maze, and in the light compartment of a light–dark box. In most cases, the effects of cocaine SA were dependent on the pattern/amount of cocaine intake with statistically significant differences from saline self-administering controls only observed in LgA rats. Conclusions When examined after several weeks of extinction/ withdrawal, cocaine SA promotes a more active pattern of behavior during times of stress that is associated with a heightened susceptibility to stressor-induced cocaine-seeking behavior and may be the consequence of augmented CRF regulation of addiction-related neurocircuitry

Pathogen Recognition Receptor Signaling Accelerates Phosphorylation-Dependent Degradation of IFNAR1

An ability to sense pathogens by a number of specialized cell types including the dendritic cells plays a central role in host's defenses. Activation of these cells through the stimulation of the pathogen-recognition receptors induces the production of a number of cytokines including Type I interferons (IFNs) that mediate the diverse mechanisms of innate immunity. Type I IFNs interact with the Type I IFN receptor, composed of IFNAR1 and IFNAR2 chains, to mount the host defense responses. However, at the same time, Type I IFNs elicit potent anti-proliferative and pro-apoptotic effects that could be detrimental for IFN-producing cells. Here, we report that the activation of p38 kinase in response to pathogen-recognition receptors stimulation results in a series of phosphorylation events within the IFNAR1 chain of the Type I IFN receptor. This phosphorylation promotes IFNAR1 ubiquitination and accelerates the proteolytic turnover of this receptor leading to an attenuation of Type I IFN signaling and the protection of activated dendritic cells from the cytotoxic effects of autocrine or paracrine Type I IFN. In this paper we discuss a potential role of this mechanism in regulating the processes of innate immunity

Endocytosis of the IFNAR1 chain of Type 1 interferon receptor is regulated by diverse E2 ubiquitin conjugation enzymes

Ubiquitination of signaling receptors triggers their endocytosis to restrict the extent of cell signaling. Type 1 interferon (IFN1) eliminates its receptor from cell surface via stimulating the ubiquitination of its IFNAR1 chain. While it was suggested that this ubiquitination aids IFNAR1internalization via relieving a steric hindrance of a linear motif within IFNAR1 from the endocytic machinery, the mechanisms involved remain poorly understood. Here we describe a specific role for two disparate ubiquitin acceptor sites within this receptor. These sites, Lys501 and Lys525 / 526, exhibit a preference for polyubiquitination via either Lys63- or Lys48‑linked chains (K63‑Ub and K48‑Ub, respectively). Whereas the SCFβTrcp E3 ubiquitin ligase controls either type of ubiquitination-dependent IFNAR1 endocytosis, the specificity of these processes is determined by two different E2 ubiquitin conjugating enzymes, Ubc13 and Cdc34. These enzymes can be directly used by SCFβTrcp E3 ubiquitin ligase to generate either K63‑Ub or K48‑Ub in vitro. Ubc13 is involved in IFNAR1 endocytosis driven by the K63‑Ub modification of Lys501, whereas the K48‑Ub-specific Cdc34 affects receptor endocytosis via ubiquitin conjugation that occurs on Lys525 / 526. Both types of linkages combine to maximize IFNAR1 endocytosis otherwise suppressed by unfavorable conformation dependent on the presence of a conserved Pro470 within the intracellular domain of IFNAR1. We propose a model where alternate utilization of both E2s to assemble diverse polyubiquitin linkages cooperates to achieve IFNAR1 intracellular domain conformations and spatial arrangements that favor a maximal rate of receptor endocytosis