Dataset Information

Proteomic survey of ubiquitin-linked nuclear proteins in interferon-stimulated macrophages.

ABSTRACT: Ubiquitin modification plays a critical role in immune responses. Some cytoplasmic factors require ubiquitination to execute proper signaling upon pathogen and cytokine stimulation. However, ubiquitin modification and its functional significance have not been fully studied for many nuclear proteins. We report here that stimulation of RAW macrophages with interferon-γ and toll-like receptor ligands that activates innate immune responses triggers a global increase in ubiquitinated proteins in the nucleus, pointing to the role for ubiquitin modification in regulating nuclear events during innate immune responses. By immunopurification and mass-spectrometry analyses, we found that more than 200 proteins are directly or indirectly associated with ubiquitin in stimulated RAW cells. These proteins included proteins in the ubiquitin pathways, those involved in DNA metabolism, chromatin and transcriptional regulation, and mRNA processing. The largest group of proteins found in our list was ribosomal proteins important for protein translation. Other proteins found here were heat shock proteins and stress-response factors, suggesting a link between macrophage activation and stress response. In conclusion, upon macrophage activation, a large number of nuclear proteins become associated with ubiquitin modification, presumably leading to a global shift in the genome activity, important for proper execution of innate immune responses.

SUBMITTER: Kim JY

PROVIDER: S-EPMC3151623 | biostudies-literature | 2011 Aug

REPOSITORIES: biostudies-literature

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Proteomic survey of ubiquitin-linked nuclear proteins in interferon-stimulated macrophages.

Kim Ji Young JY Anderson Eric D ED Huynh Walter W Dey Anup A Ozato Keiko K

Journal of interferon & cytokine research : the official journal of the International Society for Interferon and Cytokine Research 20110323 8

Ubiquitin modification plays a critical role in immune responses. Some cytoplasmic factors require ubiquitination to execute proper signaling upon pathogen and cytokine stimulation. However, ubiquitin modification and its functional significance have not been fully studied for many nuclear proteins. We report here that stimulation of RAW macrophages with interferon-γ and toll-like receptor ligands that activates innate immune responses triggers a global increase in ubiquitinated proteins in the ...[more]

PMID: 21428739

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Project description:Viruses manipulate the complex interferon and interferon-stimulated gene (ISG) system in different ways. We have previously shown that HIV inhibits type I and III interferons in its key target cells but directly stimulates a subset of >20 ISGs in macrophages and dendritic cells, many of which are antiviral. Here, we examine the mechanism of induction of ISGs and show this occurs in two phases. The first phase was transient (0 to 24 h postinfection [hpi]), induced mainly by extracellular vesicles and one of its component proteins, HSP90α, contained within the HIV inoculum. The second, dominant, and persistent phase (>48 hpi) was induced via newly transcribed HIV RNA and sensed via RIGI, as shown by the reduction in ISG expression after the knockdown of the RIGI adaptor, MAVS, by small interfering RNA (siRNA) and the inhibition of both the initiation and elongation of HIV transcription by short hairpin RNA (shRNA) transcriptional silencing. We further define the induction pathway, showing sequential HIV RNA stimulation via Tat, RIGI, MAVS, IRF1, and IRF7, also identified by siRNA knockdown. IRF1 also plays a key role in the first phase. We also show that the ISGs IFIT1 to -3 inhibit HIV production, measured as extracellular infectious virus. All induced antiviral ISGs probably lead to restriction of HIV replication in macrophages, contributing to a persistent, noncytopathic infection, while the inhibition of interferon facilitates spread to adjacent cells. Both may influence the size of macrophage HIV reservoirs in vivo Elucidating the mechanisms of ISG induction may help in devising immunotherapeutic strategies to limit the size of these reservoirs.IMPORTANCE HIV, like other viruses, manipulates the antiviral interferon and interferon-stimulated gene (ISG) system to facilitate its initial infection and establishment of viral reservoirs. HIV specifically inhibits all type I and III interferons in its target cells, including macrophages, dendritic cells, and T cells. It also induces a subset of over 20 ISGs of differing compositions in each cell target. This occurs in two temporal phases in macrophages. Extracellular vesicles contained within the inoculum induce the first, transient phase of ISGs. Newly transcribed HIV RNA induce the second, dominant ISG phase, and here, the full induction pathway is defined. Therefore, HIV nucleic acids, which are potent inducers of interferon and ISGs, are initially concealed, and antiviral ISGs are not fully induced until replication is well established. These antiviral ISGs may contribute to persistent infection in macrophages and to the establishment of viral reservoirs in vivo.

Dataset Information

Proteomic survey of ubiquitin-linked nuclear proteins in interferon-stimulated macrophages.

Publications

Proteomic survey of ubiquitin-linked nuclear proteins in interferon-stimulated macrophages.

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