Dataset Information

DNA Polymerase ? Interacts with Both the TRAF-like and UBL1-2 Domains of USP7.

ABSTRACT: Reversible protein ubiquitination is an essential signaling mechanism within eukaryotes. Deubiquitinating enzymes are critical to this process, as they mediate removal of ubiquitin from substrate proteins. Ubiquitin-specific protease 7 (USP7) is a prominent deubiquitinating enzyme, with an extensive network of interacting partners and established roles in cell cycle activation, immune responses and DNA replication. Characterized USP7 substrates primarily interact with one of two major binding sites outside the catalytic domain. These are located on the USP7 N-terminal TRAF-like (TRAF) domain and the first and second UBL domains (UBL1-2) within the C-terminal tail. Here, we report that DNA polymerase iota (Pol ?) is a novel USP7 substrate that interacts with both TRAF and UBL1-2. Through the use of biophysical approaches and mutational analysis, we characterize both interfaces and demonstrate that bipartite binding to both USP7 domains is required for efficient Pol ? deubiquitination. Together, these data establish a new bipartite mode of USP7 substrate binding.

SUBMITTER: Ashton NW

PROVIDER: S-EPMC7873624 | biostudies-literature | 2021 Jan

REPOSITORIES: biostudies-literature

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DNA Polymerase ι Interacts with Both the TRAF-like and UBL1-2 Domains of USP7.

Ashton Nicholas W NW Ashton Nicholas W NW Valles Gabrielle J GJ Jaiswal Nancy N Bezsonova Irina I Woodgate Roger R

Journal of molecular biology 20201203 2

Reversible protein ubiquitination is an essential signaling mechanism within eukaryotes. Deubiquitinating enzymes are critical to this process, as they mediate removal of ubiquitin from substrate proteins. Ubiquitin-specific protease 7 (USP7) is a prominent deubiquitinating enzyme, with an extensive network of interacting partners and established roles in cell cycle activation, immune responses and DNA replication. Characterized USP7 substrates primarily interact with one of two major binding si ...[more]

PMID: 33279577

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Project description:Human herpesvirus 8 (HHV-8) encodes four viral interferon regulatory factors (vIRFs 1 to 4), all of which are expressed during lytic replication and inhibit a variety of antiviral signaling pathways. Viral IRFs 1, 2, and 3 are also expressed during latency in primary effusion lymphoma (PEL) cells, and vIRF-1 and vIRF-3 have been reported to promote PEL cell viability. Viral IRFs 1, 3, and 4 are known to interact with ubiquitin-specific protease 7 (USP7); interactions of vIRF-1 and vIRF-3 with USP7 promote PEL cell viability and regulate productive replication. Here, we report that vIRF-2 also targets USP7, utilizing a PSTS motif matching the USP7 N-terminal domain-binding A/PxxS consensus, but uniquely requires catalytic domain residues for intracellular interaction. In functional and mechanistic analyses, tumor necrosis factor receptor-associated factor (TRAF)-mediated signaling and associated polyubiquitination of TRAFs 3 and 6, specifically, were regulated negatively by USP7 and positively by vIRF-2-USP7 interaction, the latter competing for USP7-TRAF association. Using depletion, depletion-complementation, and targeted mutagenesis approaches, vIRF-2 was determined to promote latent PEL cell viability, likely independently of USP7 interaction, while lytic replication was inhibited by vIRF-2, in part or in whole via USP7 interaction. Together, our data identify a new molecular determinant of USP7 recognition, TRAF3/6-specific targeting by the deubiquitinase, associated activation of these TRAFs by vIRF-2, and activities of vIRF-2 and vIRF-2-USP7 interaction in HHV-8 latent and lytic biology.IMPORTANCE Human herpesvirus 8-encoded IRF homologues were the first to be identified in a virus. Through inhibitory interactions with cellular IRFs and other mediators of antiviral signaling, the vIRFs are believed to be essential for productive replication and also for latency in particular cell types. The deubiquitinase USP7 is a regulator of key cellular pathways, modulates HHV-8 latent and lytic infection, and is targeted by vIRFs 1, 3, and 4. Here, we report that vIRF-2 also interacts with USP7, via a means distinguishable from USP7 interactions with other vIRFs and other proteins, that this interaction modulates antiviral signaling via disruption of USP7 interactions with innate immune signaling proteins TRAF3 and TRAF6, and that vIRF-2 targeting of USP7 regulates HHV-8 productive replication. The presented data are the first to identify vIRF-2 targeting of USP7 and its role in HHV-8 biology, expanding our understanding of the repertoire and importance of virus-host interactions.

Dataset Information

DNA Polymerase ? Interacts with Both the TRAF-like and UBL1-2 Domains of USP7.

Publications

DNA Polymerase ι Interacts with Both the TRAF-like and UBL1-2 Domains of USP7.

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