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LUBAC synthesizes linear ubiquitin chains via a thioester intermediate.

ABSTRACT: The linear ubiquitin chain assembly complex (LUBAC) is a RING E3 ligase that regulates immune and inflammatory signalling pathways. Unlike classical RING E3 ligases, LUBAC determines the type of ubiquitin chain being formed, an activity normally associated with the E2 enzyme. We show that the RING-in-between-RING (RBR)-containing region of HOIP--the catalytic subunit of LUBAC--is sufficient to generate linear ubiquitin chains. However, this activity is inhibited by the N-terminal portion of the molecule, an inhibition that is released upon complex formation with HOIL-1L or SHARPIN. Furthermore, we demonstrate that HOIP transfers ubiquitin to the substrate through a thioester intermediate formed by a conserved cysteine in the RING2 domain, supporting the notion that RBR ligases act as RING/HECT hybrids.

SUBMITTER: Stieglitz B

PROVIDER: S-EPMC3432797 | biostudies-literature | 2012 Sep

REPOSITORIES: biostudies-literature

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LUBAC synthesizes linear ubiquitin chains via a thioester intermediate.

Stieglitz Benjamin B Morris-Davies Aylin C AC Koliopoulos Marios G MG Christodoulou Evangelos E Rittinger Katrin K

EMBO reports 20120713 9

The linear ubiquitin chain assembly complex (LUBAC) is a RING E3 ligase that regulates immune and inflammatory signalling pathways. Unlike classical RING E3 ligases, LUBAC determines the type of ubiquitin chain being formed, an activity normally associated with the E2 enzyme. We show that the RING-in-between-RING (RBR)-containing region of HOIP--the catalytic subunit of LUBAC--is sufficient to generate linear ubiquitin chains. However, this activity is inhibited by the N-terminal portion of the ...[more]

PMID: 22791023

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The linear ubiquitin chain assembly complex (LUBAC) generates heterotypic ubiquitin chains.

Project description:The linear ubiquitin chain assembly complex (LUBAC) is the only known ubiquitin ligase for linear/Met1-linked ubiquitin chain formation. One of the LUBAC components, heme-oxidized IRP2 ubiquitin ligase 1 (HOIL-1L), was recently shown to catalyse oxyester bond formation between ubiquitin and some substrates. However, oxyester bond formation in the context of LUBAC has not been directly observed. Here, we present the first 3D reconstruction of human LUBAC obtained by electron microscopy and report its generation of heterotypic ubiquitin chains containing linear linkages with oxyester-linked branches. We found that this event depends on HOIL-1L catalytic activity. By cross-linking mass spectrometry showing proximity between the catalytic RING-in-between-RING (RBR) domains, a coordinated ubiquitin relay mechanism between the HOIL-1-interacting protein (HOIP) and HOIL-1L ligases is suggested. In mouse embryonic fibroblasts, these heterotypic chains were induced by TNF, which is reduced in cells expressing an HOIL-1L catalytic inactive mutant. In conclusion, we demonstrate that LUBAC assembles heterotypic ubiquitin chains by the concerted action of HOIP and HOIL-1L.

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The linear ubiquitin chain assembly complex LUBAC generates heterotypic ubiquitin chains

Project description:The linear ubiquitin chain assembly complex (LUBAC) is the only known ubiquitin ligase that generates linear/Met1-linked ubiquitin chains. One of the LUBAC components, HOIL-1L, was recently shown to catalyse oxyester bond formation between the C-terminus of ubiquitin and some substrates. However, oxyester bond formation in the context of LUBAC has not been directly observed. We present the first 3D reconstruction of LUBAC obtained by electron microscopy and report its generation of heterotypic ubiquitin chains containing linear linkages with oxyester-linked branches. We found that addition of the oxyester-bound branches depends on HOIL-1L catalytic activity. We suggest a coordinated ubiquitin relay mechanism between the HOIP and HOIL-1L ligases supported by cross-linking mass spectrometry data, which show proximity between the catalytic RBR domains. Mutations in the linear ubiquitin chain-binding NZF domain of HOIL-1L reduces chain branching confirming its role in the process. In cells, these heterotypic chains were induced by TNF. In conclusion, we demonstrate that LUBAC assembles heterotypic ubiquitin chains with linear and oxyester-linked branches by the concerted action of HOIP and HOIL-1L.

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LUBAC-synthesized linear ubiquitin chains restrict cytosol-invading bacteria by activating autophagy and NF-?B.

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A20 inhibits LUBAC-mediated NF-?B activation by binding linear polyubiquitin chains via its zinc finger 7.

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STING induces LUBAC synthesis of linear ubiquitin chains to activate NFkB

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The linear ubiquitin assembly complex (LUBAC) is essential for NLRP3 inflammasome activation.

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