Murine cytomegaloviruses m139 targets DDX3 to curtail interferon production and promote viral replication

Puhach, Olha; Ostermann, Eléonore; Frascaroli, Giada; Schlüter, Hartmut; Brinkmann, Melanie M.; Brune, Wolfram

doi:10.1101/2020.04.17.046516

Cited by 4 publications

(1 citation statement)

References 66 publications

(91 reference statements)

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“…CRISPR/Cas9 gene editing was used to generate Sel1L knockout MEFs essentially as described (20,42). Briefly, two different guide RNAs (5'-GTCGTTGCTGCTGCTCTGCG-3' and 5'-GCTGCTCTGCGCGGTGCTCC-3') targeting the murine Sel1L gene were designed using E-CRISP (http://www.e-crisp.org/E-CRISP/designcrispr.html) and inserted into the lentiviral vector pSicoR-CRISPR-puroR (kindly provided by R. J. Lebbink, University Medical Center Utrecht, Netherlands).…”

Section: Crispr/cas9 Gene Editingmentioning

confidence: 99%

Viral-Mediated Tethering to SEL1L Facilitates Endoplasmic Reticulum-Associated Degradation of IRE1

Hinte

Müller

Brune

2021

J Virol

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The unfolded protein response (UPR) and endoplasmic reticulum (ER)-associated degradation (ERAD) are two essential components of the quality control system for proteins in the secretory pathway. When unfolded proteins accumulate in the ER, UPR sensors such as IRE1 induce the expression of ERAD genes, thereby increasing protein export from the ER to the cytosol and subsequent degradation by the proteasome. Conversely, IRE1 itself is an ERAD substrate, indicating that the UPR and ERAD regulate each other. Viruses are intracellular parasites that exploit the host cell for their own benefit. Cytomegaloviruses selectively modulate the UPR to take advantage of beneficial and inhibit detrimental effects on viral replication. We have previously shown that murine and human cytomegaloviruses express homologous proteins (M50 and UL50, respectively) that dampen the UPR at late times post infection by inducing IRE1 degradation. However, the degradation mechanism has remained uncertain. Here we show that the cytomegalovirus M50 protein mediates IRE1 degradation by the proteasome. M50-dependent IRE1 degradation can be blocked by pharmacological inhibition of p97/VCP or by genetic ablation of SEL1L, both of which are components of the ERAD machinery. SEL1L acts as a cofactor of the E3 ubiquitin ligase HRD1, while p97/VCP is responsible for the extraction of ubiquitylated proteins from the ER to the cytosol. We further show that M50 facilitates the IRE1-SEL1L interaction by binding to both, IRE1 and SEL1L. These results indicate that the viral M50 protein dampens the UPR by tethering IRE1 to SEL1L, thereby promoting its degradation by the ERAD machinery. IMPORTANCE Viruses infect cells of their host and force them to produce virus progeny. This can impose stress on the host cell and activate counter-regulatory mechanisms. Protein overload in the endoplasmic reticulum (ER) leads to ER stress and triggers the unfolded protein response, which in turn upregulates protein folding and increases the degradation of proteins in the ER. Previous work has shown that cytomegaloviruses interfere with the unfolded protein response by degrading the sensor molecule IRE1. Herein we demonstrate how the cytomegalovirus M50 protein exploits the ER-associated degradation machinery to dispose of IRE1. Degradation of IRE1 curbs the unfolded protein response and helps the virus to increase the synthesis of its own proteins and the production of virus progeny.

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Section: Crispr/cas9 Gene Editingmentioning

confidence: 99%

Viral-Mediated Tethering to SEL1L Facilitates Endoplasmic Reticulum-Associated Degradation of IRE1

Hinte

Müller

Brune

2021

J Virol

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Viral mediated tethering to SEL1L facilitates ER-associated degradation of IRE1

Hinte

Müller

Brune

2020

Preprint

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The unfolded protein response (UPR) and endoplasmic reticulum (ER)-associated degradation (ERAD) are two essential components of the quality control system for proteins in the secretory pathway. When unfolded proteins accumulate in the ER, UPR sensors such as IRE1 induce the expression of ERAD genes, thereby increasing protein export from the ER to the cytosol and subsequent degradation by the proteasome. Conversely, IRE1 itself is an ERAD substrate, indicating that the UPR and ERAD regulate each other. Viruses are intracellular parasites that exploit the host cell for their own benefit. Cytomegaloviruses selectively modulate the UPR to take advantage of beneficial and inhibit detrimental effects on viral replication. We have previously shown that murine and human cytomegaloviruses express homologous proteins (M50 and UL50, respectively) that dampen the UPR at late times post infection by inducing IRE1 degradation. However, the degradation mechanism has remained uncertain. Here we show that the cytomegalovirus M50 protein mediates IRE1 degradation by the proteasome. M50-dependent IRE1 degradation can be blocked by pharmacological inhibition of p97/VCP or by genetic ablation of SEL1L, both of which are component of the ERAD machinery. SEL1L acts as a cofactor of the E3 ubiquitin ligase HRD1, while p97/VCP is responsible for the extraction of ubiquitylated proteins from the ER to the cytosol. We further show that M50 facilitates the IRE1-SEL1L interaction by binding to both, IRE1 and SEL1L. These results indicate that the viral M50 protein dampens the UPR by tethering IRE1 to SEL1L, thereby promoting its degradation by the ERAD machinery.ImportanceViruses infect cells of their host and force them to produce virus progeny. This can impose stress on the host cell and activate counter-regulatory mechanisms. Protein overload in the endoplasmic reticulum (ER) leads to ER stress and triggers the unfolded protein response, which in turn upregulates protein folding and increases the degradation of proteins in the ER. Previous work has shown that cytomegaloviruses interfere with the unfolded protein response by degrading the sensor molecule IRE1. Herein we demonstrate how the viral M50 protein exploits the ER-associated degradation machinery to dispose of IRE1. Degradation of IRE1 curbs the unfolded protein response and helps the virus to increase the synthesis of its own proteins.

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The impact of murine cytomegalovirus (mCMV) route and age at infection upon virus spread, immune responses and the establishment of latency

Coplen

Jergović

Terner

et al. 2021

Preprint

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Cytomegalovirus (CMV) is a ubiquitous human virus, which establishes a characteristic lifetime infection in its host. Murine CMV (mCMV) is a widely-used infection model that has been employed to investigate the nature and extent of CMV’s contribution to inflammatory, immunological, and health disturbances in humans. In an effort to assess the role of route and age in modeling hCMV infection in mice, we have performed a comparative analysis of two common experimental modes of infection (intraperitoneal and intranasal) at two different clinically relevant ages (4 weeks, or prepubescent childhood equivalent, and 12 weeks, or young postpubescent adult). We found that while both routes of infection led to similar early viral loads, differential activation of several parameters of innate immune function were observed. In particular, younger, prepubescent mice exhibited the strongest NK activation in the blood in response to i.p. infection, with this trend holding true in NK cells expressing the mCMV-specific receptor Ly49H. Moreover, i.p. infected animals accumulated a larger amount of anti-mCMV IgG and experienced a greater expansion of both acute and latent phase CD8+ T cells. This was especially true for young postpubescent mice, further illustrating a distinction in the bloodborne immune response across not only infection routes, but also ages. These results may be important in the understanding of how a more physiologically applicable model of CMV influences immunity, inflammation, and health over the lifespan.

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Murine cytomegaloviruses m139 targets DDX3 to curtail interferon production and promote viral replication

Cited by 4 publications

References 66 publications

Viral-Mediated Tethering to SEL1L Facilitates Endoplasmic Reticulum-Associated Degradation of IRE1

Viral-Mediated Tethering to SEL1L Facilitates Endoplasmic Reticulum-Associated Degradation of IRE1

Viral mediated tethering to SEL1L facilitates ER-associated degradation of IRE1

The impact of murine cytomegalovirus (mCMV) route and age at infection upon virus spread, immune responses and the establishment of latency

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