This page has only limited features, please log in for full access.

Dr. Craig Mccormick
Department of Microbiology and Immunology, Dalhousie University, Sir Charles Tupper Medical Building, Room 7-P 5850 College Street, Halifax, Nova Scotia, Canada B3H 4R2

Basic Info


Research Keywords & Expertise

0 Autophagy
0 Inflammation
0 stress granules
0 Influenza A virus
0 unfolded protein response

Fingerprints

Influenza A virus
Host shutoff
kaposi’s sarcoma-associated herpesvirus
Autophagy
stress granules
unfolded protein response
Inflammation
MRNA turnover and translation

Honors and Awards

The user has no records in this section


Career Timeline

The user has no records in this section.


Short Biography

The user biography is not available.
Following
Followers
Co Authors
The list of users this user is following is empty.
Following: 0 users

Feed

Journal article
Published: 24 March 2021 in Journal of Virology
Reads 0
Downloads 0

Influenza A viruses (IAVs) utilize host shutoff mechanisms to limit antiviral gene expression and redirect translation machinery to the synthesis of viral proteins. Previously, we showed that IAV replication is sensitive to protein synthesis inhibitors that block translation initiation and induce the formation of cytoplasmic condensates of untranslated messenger ribonucleoprotein complexes called stress granules (SGs). In this study, using an image-based high-content screen, we identified two thiopurines, 6-thioguanine (6-TG) and 6-thioguanosine (6-TGo), that triggered SG formation in IAV-infected cells and blocked IAV replication in a dose-dependent manner without eliciting SG formation in uninfected cells. 6-TG and 6-TGo selectively disrupted the synthesis and maturation of the IAV glycoproteins hemagglutinin (HA) and neuraminidase (NA) without affecting the levels of the viral RNAs that encode them. By contrast, these thiopurines had a minimal effect on other IAV proteins or global host protein synthesis. Disruption of IAV glycoprotein accumulation by 6-TG and 6-TGo correlated with the activation of the unfolded protein response (UPR) sensors activating transcription factor 6 (ATF6), inositol-requiring enzyme 1 (IRE1), and double-stranded RNA-activated protein kinase (PKR)-like endoplasmic reticulum (ER) kinase (PERK), leading to downstream UPR gene expression. Treatment of infected cells with the chemical chaperone 4-phenylbutyric acid diminished thiopurine-induced UPR activation and partially restored the processing and accumulation of HA and NA. By contrast, chemical inhibition of the integrated stress response downstream of PERK restored the accumulation of NA monomers but did not restore the processing of viral glycoproteins. Genetic deletion of PERK enhanced the antiviral effect of 6-TG without causing overt cytotoxicity, suggesting that while UPR activation correlates with diminished viral glycoprotein accumulation, PERK could limit the antiviral effects of drug-induced ER stress. Taken together, these data indicate that 6-TG and 6-TGo are effective host-targeted antivirals that trigger the UPR and selectively disrupt the accumulation of viral glycoproteins. IMPORTANCE Secreted and transmembrane proteins are synthesized in the endoplasmic reticulum (ER), where they are folded and modified prior to transport. Many viruses rely on the ER for the synthesis and processing of viral glycoproteins that will ultimately be incorporated into viral envelopes. Viral burden on the ER can trigger the unfolded protein response (UPR). Much remains to be learned about how viruses coopt the UPR to ensure efficient synthesis of viral glycoproteins. Here, we show that two FDA-approved thiopurine drugs, 6-TG and 6-TGo, induce the UPR, which represents a previously unrecognized effect of these drugs on cell physiology. This thiopurine-mediated UPR activation blocks influenza virus replication by impeding viral glycoprotein accumulation. Our findings suggest that 6-TG and 6-TGo may have broad antiviral effects against enveloped viruses that require precise tuning of the UPR to support viral glycoprotein synthesis.

ACS Style

Patrick D. Slaine; Mariel Kleer; Brett A. Duguay; Eric S. Pringle; Eileigh Kadijk; Shan Ying; Aruna Balgi; Michel Roberge; Craig McCormick; Denys A. Khaperskyy. Thiopurines Activate an Antiviral Unfolded Protein Response That Blocks Influenza A Virus Glycoprotein Accumulation. Journal of Virology 2021, 95, 1 .

AMA Style

Patrick D. Slaine, Mariel Kleer, Brett A. Duguay, Eric S. Pringle, Eileigh Kadijk, Shan Ying, Aruna Balgi, Michel Roberge, Craig McCormick, Denys A. Khaperskyy. Thiopurines Activate an Antiviral Unfolded Protein Response That Blocks Influenza A Virus Glycoprotein Accumulation. Journal of Virology. 2021; 95 (11):1.

Chicago/Turabian Style

Patrick D. Slaine; Mariel Kleer; Brett A. Duguay; Eric S. Pringle; Eileigh Kadijk; Shan Ying; Aruna Balgi; Michel Roberge; Craig McCormick; Denys A. Khaperskyy. 2021. "Thiopurines Activate an Antiviral Unfolded Protein Response That Blocks Influenza A Virus Glycoprotein Accumulation." Journal of Virology 95, no. 11: 1.

Journal article
Published: 17 November 2020 in Cell Death & Disease
Reads 0
Downloads 0

Glioblastoma (GBM) is the most common primary malignant brain tumor, and it has a uniformly poor prognosis. Hypoxia is a feature of the GBM microenvironment, and previous work has shown that cancer cells residing in hypoxic regions resist treatment. Hypoxia can trigger the formation of stress granules (SGs), sites of mRNA triage that promote cell survival. A screen of 1120 FDA-approved drugs identified 129 candidates that delayed the dissolution of hypoxia-induced SGs following a return to normoxia. Amongst these candidates, the selective estrogen receptor modulator (SERM) raloxifene delayed SG dissolution in a dose-dependent manner. SG dissolution typically occurs by 15 min post-hypoxia, however pre-treatment of immortalized U251 and U3024 primary GBM cells with raloxifene prevented SG dissolution for up to 2 h. During this raloxifene-induced delay in SG dissolution, translational silencing was sustained, eIF2α remained phosphorylated and mTOR remained inactive. Despite its well-described role as a SERM, raloxifene-mediated delay in SG dissolution was unaffected by co-administration of β-estradiol, nor did β-estradiol alone have any effect on SGs. Importantly, the combination of raloxifene and hypoxia resulted in increased numbers of late apoptotic/necrotic cells. Raloxifene and hypoxia also demonstrated a block in late autophagy similar to the known autophagy inhibitor chloroquine (CQ). Genetic disruption of the SG-nucleating proteins G3BP1 and G3BP2 revealed that G3BP1 is required to sustain the raloxifene-mediated delay in SG dissolution. Together, these findings indicate that modulating the stress response can be used to exploit the hypoxic niche of GBM tumors, causing cell death by disrupting pro-survival stress responses and control of protein synthesis.

ACS Style

Kathleen M. Attwood; Aaron Robichaud; Lauren P. Westhaver; Elizabeth L. Castle; David M. Brandman; Aruna D. Balgi; Michel Roberge; Patricia Colp; Sidney Croul; Inhwa Kim; Craig McCormick; Jennifer A. Corcoran; Adrienne Weeks. Raloxifene prevents stress granule dissolution, impairs translational control and promotes cell death during hypoxia in glioblastoma cells. Cell Death & Disease 2020, 11, 989 .

AMA Style

Kathleen M. Attwood, Aaron Robichaud, Lauren P. Westhaver, Elizabeth L. Castle, David M. Brandman, Aruna D. Balgi, Michel Roberge, Patricia Colp, Sidney Croul, Inhwa Kim, Craig McCormick, Jennifer A. Corcoran, Adrienne Weeks. Raloxifene prevents stress granule dissolution, impairs translational control and promotes cell death during hypoxia in glioblastoma cells. Cell Death & Disease. 2020; 11 (11):989.

Chicago/Turabian Style

Kathleen M. Attwood; Aaron Robichaud; Lauren P. Westhaver; Elizabeth L. Castle; David M. Brandman; Aruna D. Balgi; Michel Roberge; Patricia Colp; Sidney Croul; Inhwa Kim; Craig McCormick; Jennifer A. Corcoran; Adrienne Weeks. 2020. "Raloxifene prevents stress granule dissolution, impairs translational control and promotes cell death during hypoxia in glioblastoma cells." Cell Death & Disease 11, no. 11: 989.

Preprint content
Published: 01 October 2020
Reads 0
Downloads 0

Enveloped viruses, including influenza A viruses (IAVs) and coronaviruses (CoVs), utilize the host cell secretory pathway to synthesize viral glycoproteins and direct them to sites of assembly. Using an image-based high-content screen, we identified two thiopurines, 6-thioguanine (6-TG) and 6-thioguanosine (6-TGo), that selectively disrupted the processing and accumulation of IAV glycoproteins hemagglutinin (HA) and neuraminidase (NA). Selective disruption of IAV glycoprotein processing and accumulation by 6-TG and 6-TGo correlated with unfolded protein response (UPR) activation and HA accumulation could be partially restored by the chemical chaperone 4-phenylbutyrate (4PBA). Chemical inhibition of the integrated stress response (ISR) restored accumulation of NA monomers in the presence of 6-TG or 6-TGo, but did not restore NA glycosylation or oligomerization. Thiopurines inhibited replication of the human coronavirus OC43 (HCoV-OC43), which also correlated with UPR/ISR activation and diminished accumulation of ORF1ab and nucleocapsid (N) mRNAs and N protein, which suggests broader disruption of coronavirus gene expression in ER-derived cytoplasmic compartments. The chemically similar thiopurine 6-mercaptopurine (6-MP) had little effect on the UPR and did not affect IAV or HCoV-OC43 replication. Consistent with reports on other CoV Spike (S) proteins, ectopic expression of SARS-CoV-2 S protein caused UPR activation. 6-TG treatment inhibited accumulation of full length S0 or furin-cleaved S2 fusion proteins, but spared the S1 ectodomain. DBeQ, which inhibits the p97 AAA-ATPase required for retrotranslocation of ubiquitinated misfolded proteins during ER-associated degradation (ERAD) restored accumulation of S0 and S2 proteins in the presence of 6-TG, suggesting that 6-TG induced UPR accelerates ERAD-mediated turnover of membrane-anchored S0 and S2 glycoproteins. Taken together, these data indicate that 6-TG and 6-TGo are effective host-targeted antivirals that trigger the UPR and disrupt accumulation of viral glycoproteins. Importantly, our data demonstrate for the first time the efficacy of these thiopurines in limiting IAV and HCoV-OC43 replication in cell culture models. IMPORTANCE Secreted and transmembrane proteins are synthesized in the endoplasmic reticulum (ER), where they are folded and modified prior to transport. During infection, many viruses burden the ER with the task of creating and processing viral glycoproteins that will ultimately be incorporated into viral envelopes. Some viruses refashion the ER into replication compartments where viral gene expression and genome replication take place. This viral burden on the ER can trigger the cellular unfolded protein response (UPR), which attempts to increase the protein folding and processing capacity of the ER to match the protein load. Much remains to be learned about how viruses co-opt the UPR to ensure efficient synthesis of viral glycoproteins. Here, we show that two FDA-approved thiopurine drugs, 6-TG and 6-TGo, induce the UPR in a manner that impedes viral glycoprotein accumulation for enveloped influenza viruses and coronaviruses. These drugs may impede the replication of viruses that require precise tuning of the UPR to support viral glycoprotein synthesis for the successful completion of a replication cycle.

ACS Style

Patrick D Slaine; Mariel Kleer; Brett Duguay; Eric Stanley Pringle; Eileigh Kadijk; Shan Ying; Aruna D. Balgi; Michel Roberge; Craig McCormick; Denys A. Khaperskyy. Thiopurines activate an antiviral unfolded protein response that blocks viral glycoprotein accumulation in cell culture infection model. 2020, 1 .

AMA Style

Patrick D Slaine, Mariel Kleer, Brett Duguay, Eric Stanley Pringle, Eileigh Kadijk, Shan Ying, Aruna D. Balgi, Michel Roberge, Craig McCormick, Denys A. Khaperskyy. Thiopurines activate an antiviral unfolded protein response that blocks viral glycoprotein accumulation in cell culture infection model. . 2020; ():1.

Chicago/Turabian Style

Patrick D Slaine; Mariel Kleer; Brett Duguay; Eric Stanley Pringle; Eileigh Kadijk; Shan Ying; Aruna D. Balgi; Michel Roberge; Craig McCormick; Denys A. Khaperskyy. 2020. "Thiopurines activate an antiviral unfolded protein response that blocks viral glycoprotein accumulation in cell culture infection model." , no. : 1.

Review
Published: 14 July 2020 in Viruses
Reads 0
Downloads 0

Basic leucine zipper (bZIP) transcription factors (TFs) govern diverse cellular processes and cell fate decisions. The hallmark of the leucine zipper domain is the heptad repeat, with leucine residues at every seventh position in the domain. These leucine residues enable homo- and heterodimerization between ZIP domain α-helices, generating coiled-coil structures that stabilize interactions between adjacent DNA-binding domains and target DNA substrates. Several cancer-causing viruses encode viral bZIP TFs, including human T-cell leukemia virus (HTLV), hepatitis C virus (HCV) and the herpesviruses Marek’s disease virus (MDV), Epstein–Barr virus (EBV) and Kaposi’s sarcoma-associated herpesvirus (KSHV). Here, we provide a comprehensive review of these viral bZIP TFs and their impact on viral replication, host cell responses and cell fate.

ACS Style

Madeleine L. Stolz; Craig McCormick. The bZIP Proteins of Oncogenic Viruses. Viruses 2020, 12, 757 .

AMA Style

Madeleine L. Stolz, Craig McCormick. The bZIP Proteins of Oncogenic Viruses. Viruses. 2020; 12 (7):757.

Chicago/Turabian Style

Madeleine L. Stolz; Craig McCormick. 2020. "The bZIP Proteins of Oncogenic Viruses." Viruses 12, no. 7: 757.

Paper
Published: 24 June 2020 in Environmental Science: Water Research & Technology
Reads 0
Downloads 0

A commercially-available UV disinfection system used for hospital room disinfection was characterized and used for N95 filtering facepiece respirator (FFR) material disinfection.

ACS Style

C. Carolina Ontiveros; Crystal L. Sweeney; Christopher Smith; Sean MacIsaac; Sebastian Munoz; Ross Davidson; Craig McCormick; Nikhil Thomas; Ian Davis; Amina K. Stoddart; Graham A. Gagnon. Characterization of a commercially-available, low-pressure UV lamp as a disinfection system for decontamination of common nosocomial pathogens on N95 filtering facepiece respirator (FFR) material. Environmental Science: Water Research & Technology 2020, 6, 2089 -2102.

AMA Style

C. Carolina Ontiveros, Crystal L. Sweeney, Christopher Smith, Sean MacIsaac, Sebastian Munoz, Ross Davidson, Craig McCormick, Nikhil Thomas, Ian Davis, Amina K. Stoddart, Graham A. Gagnon. Characterization of a commercially-available, low-pressure UV lamp as a disinfection system for decontamination of common nosocomial pathogens on N95 filtering facepiece respirator (FFR) material. Environmental Science: Water Research & Technology. 2020; 6 (8):2089-2102.

Chicago/Turabian Style

C. Carolina Ontiveros; Crystal L. Sweeney; Christopher Smith; Sean MacIsaac; Sebastian Munoz; Ross Davidson; Craig McCormick; Nikhil Thomas; Ian Davis; Amina K. Stoddart; Graham A. Gagnon. 2020. "Characterization of a commercially-available, low-pressure UV lamp as a disinfection system for decontamination of common nosocomial pathogens on N95 filtering facepiece respirator (FFR) material." Environmental Science: Water Research & Technology 6, no. 8: 2089-2102.

Journal article
Published: 16 June 2020 in Journal of Virology
Reads 0
Downloads 0

Human leukocyte antigens (HLAs) are cell surface proteins that regulate innate and adaptive immune responses to viral infection by engaging with receptors on immune cells. Many viruses have evolved ways to evade host immune responses by modulating HLA expression and/or processing. Here, we provide evidence that aberrant RNA products of influenza virus genome replication can trigger retinoic acid-inducible gene I (RIG-I)/mitochondrial antiviral signaling (MAVS)-dependent remodeling of the cell surface, increasing surface presentation of HLA proteins known to inhibit the activation of an immune cell known as a natural killer (NK) cell. While this HLA upregulation would seem to be advantageous to the virus, it is kept in check by the viral nonstructural 1 (NS1) protein, which limits RIG-I activation and interferon production by the infected cell.

ACS Style

Mir Munir A. Rahim; Brendon D. Parsons; Emma L. Price; Patrick D. Slaine; Becca L. Chilvers; Gregory S. Seaton; Andrew Wight; Daniel Medina-Luna; Sayanti Dey; Shannen L. Grandy; Lauryn E. Anderson; Natalia Zamorano Cuervo; Nathalie Grandvaux; Marta M. Gaglia; Alyson A. Kelvin; Denys A. Khaperskyy; Craig McCormick; Andrew P. Makrigiannis. Defective Influenza A Virus RNA Products Mediate MAVS-Dependent Upregulation of Human Leukocyte Antigen Class I Proteins. Journal of Virology 2020, 94, 1 .

AMA Style

Mir Munir A. Rahim, Brendon D. Parsons, Emma L. Price, Patrick D. Slaine, Becca L. Chilvers, Gregory S. Seaton, Andrew Wight, Daniel Medina-Luna, Sayanti Dey, Shannen L. Grandy, Lauryn E. Anderson, Natalia Zamorano Cuervo, Nathalie Grandvaux, Marta M. Gaglia, Alyson A. Kelvin, Denys A. Khaperskyy, Craig McCormick, Andrew P. Makrigiannis. Defective Influenza A Virus RNA Products Mediate MAVS-Dependent Upregulation of Human Leukocyte Antigen Class I Proteins. Journal of Virology. 2020; 94 (13):1.

Chicago/Turabian Style

Mir Munir A. Rahim; Brendon D. Parsons; Emma L. Price; Patrick D. Slaine; Becca L. Chilvers; Gregory S. Seaton; Andrew Wight; Daniel Medina-Luna; Sayanti Dey; Shannen L. Grandy; Lauryn E. Anderson; Natalia Zamorano Cuervo; Nathalie Grandvaux; Marta M. Gaglia; Alyson A. Kelvin; Denys A. Khaperskyy; Craig McCormick; Andrew P. Makrigiannis. 2020. "Defective Influenza A Virus RNA Products Mediate MAVS-Dependent Upregulation of Human Leukocyte Antigen Class I Proteins." Journal of Virology 94, no. 13: 1.

Preprint content
Published: 31 January 2020
Reads 0
Downloads 0

Influenza A virus (IAV) increases presentation of class I human leukocyte antigen (HLA) proteins that limit antiviral responses mediated by natural killer (NK) cells, but molecular mechanisms have not yet been fully elucidated. We observed that infection with A/Fort Monmouth/1/1947 (H1N1) IAV significantly increased presentation of HLA-B, -C and -E on lung epithelial cells. Virus entry was not sufficient to induce HLA upregulation, because UV-inactivated virus had no effect. We found that HLA upregulation was elicited by aberrant internally-deleted viral RNAs (vRNAs) known as mini viral RNAs (mvRNAs) and defective interfering RNAs (DI RNAs), which bind to retinoic acid-inducible gene-I (RIG-I) and initiate mitochondrial antiviral signaling (MAVS) protein-dependent antiviral interferon (IFN) responses. Indeed, MAVS was required for HLA upregulation in response to IAV infection or ectopic mvRNA/DI RNA expression. The effect was partially due to paracrine signalling, as we observed that IAV infection or mvRNA/DI RNA-expression stimulated production of IFN-β and IFN-λ1, and conditioned media from these cells elicited a modest increase in HLA surface levels in naïve epithelial cells. HLA upregulation in response to aberrant viral RNAs could be prevented by chemical blockade of IFN receptor signal transduction. While HLA upregulation would seem to be advantageous to the virus, it is kept in check by the viral non-structural 1 (NS1) protein; we determined that NS1 limits cell-intrinsic and paracrine mechanisms of HLA upregulation. Taken together, our findings indicate that aberrant IAV RNAs stimulate HLA presentation, which may aid viral evasion of innate immunity.IMPORTANCEHuman leukocyte antigens (HLA) are cell surface proteins that regulate innate and adaptive immune responses to viral infection by engaging with receptors on immune cells. Many viruses have evolved ways to evade host immune responses by modulating HLA expression and/or processing. Here, we provide evidence that aberrant RNA products of influenza virus genome replication can trigger RIG-I/MAVS-dependent remodeling of the cell surface, increasing surface presentation of HLA proteins known to inhibit the activation of an immune cell known as a natural killer (NK) cell. While this HLA upregulation would seem to be advantageous to the virus, it is kept in check by the viral non-structural 1 (NS1) protein, which limits RIG-I activation and interferon production by the infected cell.

ACS Style

Mir Munir A. Rahim; Brendon D. Parsons; Emma L. Price; Patrick D. Slaine; Becca L. Chilvers; Gregory S. Seaton; Andrew Wight; Sayanti Dey; Shannen L. Grandy; Lauryn E. Anderson; Natalia Zamorano Cuervo; Nathalie Grandvaux; Marta M. Gaglia; Alyson A. Kelvin; Denys A. Khaperskyy; Craig McCormick; Andrew P. Makrigiannis. Defective influenza A virus RNA products mediate MAVS-dependent upregulation of human leukocyte antigen class I proteins. 2020, 1 .

AMA Style

Mir Munir A. Rahim, Brendon D. Parsons, Emma L. Price, Patrick D. Slaine, Becca L. Chilvers, Gregory S. Seaton, Andrew Wight, Sayanti Dey, Shannen L. Grandy, Lauryn E. Anderson, Natalia Zamorano Cuervo, Nathalie Grandvaux, Marta M. Gaglia, Alyson A. Kelvin, Denys A. Khaperskyy, Craig McCormick, Andrew P. Makrigiannis. Defective influenza A virus RNA products mediate MAVS-dependent upregulation of human leukocyte antigen class I proteins. . 2020; ():1.

Chicago/Turabian Style

Mir Munir A. Rahim; Brendon D. Parsons; Emma L. Price; Patrick D. Slaine; Becca L. Chilvers; Gregory S. Seaton; Andrew Wight; Sayanti Dey; Shannen L. Grandy; Lauryn E. Anderson; Natalia Zamorano Cuervo; Nathalie Grandvaux; Marta M. Gaglia; Alyson A. Kelvin; Denys A. Khaperskyy; Craig McCormick; Andrew P. Makrigiannis. 2020. "Defective influenza A virus RNA products mediate MAVS-dependent upregulation of human leukocyte antigen class I proteins." , no. : 1.

Abstract
Published: 01 January 2020 in Proceedings
Reads 0
Downloads 0

Herpesviruses usurp host cell protein synthesis machinery to convert viral mRNAs into proteins, and the endoplasmic reticulum (ER) to ensure the proper folding, post-translational modification and trafficking of secreted and transmembrane viral proteins. Overloading the ER folding capacity activates the unfolded protein response (UPR), whereby sensor proteins, ATF6, PERK and IRE1, initiate a stress-mitigating transcription program that accelerates the catabolism of misfolded proteins, while increasing the ER folding capacity. Kaposi’s sarcoma-associated herpesvirus (KSHV) can be reactivated from latency through the chemical induction of ER stress, which causes an accumulation of the XBP1s transcription factor that transactivates the viral RTA lytic switch gene. The presence of XBP1s-responsive elements in the RTA promoter suggests that KSHV evolved a mechanism to respond to ER stress. Here, we report that ATF6, PERK and IRE1 were activated upon reactivation from latency and were required for efficient KSHV lytic replication. The genetic or pharmacologic inhibitions of each UPR sensor reduced virion production. Despite UPR sensor activation during KSHV lytic replication, downstream UPR transcriptional responses were restricted: (1) ATF6 was cleaved to activate the ATF6(N) transcription factor but ATF6(N)-responsive genes were not transcribed; (2) PERK phosphorylated eIF2, but ATF4 did not accumulate; (3) IRE1 caused XBP1–mRNA splicing, but the XBP1s protein did not accumulate and the XBP1s-responsive genes were not transcribed. The complementation of XBP1s deficiency during KSHV lytic replication inhibited virion production in a dose-dependent manner in epithelial cells. Taken together, these findings indicate that, while XBP1s plays an important role in reactivation from latency, it can inhibit virus replication at a later step, which the virus overcomes by preventing its synthesis. These findings suggest that KSHV hijacks UPR sensors to promote efficient viral replication while sustaining ER stress.

ACS Style

Benjamin P. Johnston; Eric S. Pringle; Craig McCormick. KSHV Activates Unfolded Protein Response Sensors but Suppresses Downstream Transcriptional Responses to Support Lytic Replication. Proceedings 2020, 50, 116 .

AMA Style

Benjamin P. Johnston, Eric S. Pringle, Craig McCormick. KSHV Activates Unfolded Protein Response Sensors but Suppresses Downstream Transcriptional Responses to Support Lytic Replication. Proceedings. 2020; 50 (1):116.

Chicago/Turabian Style

Benjamin P. Johnston; Eric S. Pringle; Craig McCormick. 2020. "KSHV Activates Unfolded Protein Response Sensors but Suppresses Downstream Transcriptional Responses to Support Lytic Replication." Proceedings 50, no. 1: 116.

Abstract
Published: 01 January 2020 in Proceedings
Reads 0
Downloads 0

Herpesvirus genomes are decoded by host RNA polymerase enzymes, generating messenger ribonucleotides (mRNA) that are post-transcriptionally modified and exported to the cytoplasm through the combined work of host and viral factors. These viral mRNA bear 5′-m7GTP caps and poly(A) tails that should permit the assembly of canonical host eIF4F cap-binding complexes to initiate protein synthesis. However, the precise mechanisms of translation initiation remain to be investigated for Kaposi’s sarcoma-associated herpesvirus (KSHV) and other herpesviruses. During KSHV lytic replication in lymphoid cells, the activation of caspases leads to the cleavage of eIF4G and depletion of eIF4F. Translating mRNPs depleted of eIF4F retain viral mRNA, suggesting that non-eIF4F translation initiation is sufficient to support viral protein synthesis. To identify proteins required to support viral protein synthesis, we isolated and characterized actively translating messenger ribonucleoprotein (mRNP) complexes by ultracentrifugation and sucrose-gradient fractionation followed by quantitative mass spectrometry. The abundance of host translation initiation factors available to initiate viral protein synthesis were comparable between cells undergoing KSHV lytic or latent replication. The translation initiation factors eIF4E2, NCBP1, eIF4G2, and eIF3d were detected in association with actively translating mRNP complexes during KSHV lytic replication, but their depletion by RNA silencing did not affect virion production. By contrast, the N6-methyladenosine methyltransferase METTL3 was required for optimal late gene expression and virion production, but was dispensable for genome replication. Furthermore, we detected several KSHV proteins in actively translating mRNP complexes that had not previously been shown to play roles in viral protein synthesis. We conclude that KSHV usurps distinct host translation initiation systems during latent and lytic phases of infection.

ACS Style

Eric S. Pringle; Craig McCormick. Composition of Herpesvirus Ribonucleoprotein Complexes. Proceedings 2020, 50, 119 .

AMA Style

Eric S. Pringle, Craig McCormick. Composition of Herpesvirus Ribonucleoprotein Complexes. Proceedings. 2020; 50 (1):119.

Chicago/Turabian Style

Eric S. Pringle; Craig McCormick. 2020. "Composition of Herpesvirus Ribonucleoprotein Complexes." Proceedings 50, no. 1: 119.

Review
Published: 21 December 2019 in Viruses
Reads 0
Downloads 0

Herpesviruses usurp cellular stress responses to promote viral replication and avoid immune surveillance. The unfolded protein response (UPR) is a conserved stress response that is activated when the protein load in the ER exceeds folding capacity and misfolded proteins accumulate. The UPR aims to restore protein homeostasis through translational and transcriptional reprogramming; if homeostasis cannot be restored, the UPR switches from “helper” to “executioner”, triggering apoptosis. It is thought that the burst of herpesvirus glycoprotein synthesis during lytic replication causes ER stress, and that these viruses may have evolved mechanisms to manage UPR signaling to create an optimal niche for replication. The past decade has seen considerable progress in understanding how herpesviruses reprogram the UPR. Here we provide an overview of the molecular events of UPR activation, signaling and transcriptional outputs, and highlight key evidence that herpesviruses hijack the UPR to aid infection.

ACS Style

Benjamin P. Johnston; Craig McCormick. Herpesviruses and the Unfolded Protein Response. Viruses 2019, 12, 17 .

AMA Style

Benjamin P. Johnston, Craig McCormick. Herpesviruses and the Unfolded Protein Response. Viruses. 2019; 12 (1):17.

Chicago/Turabian Style

Benjamin P. Johnston; Craig McCormick. 2019. "Herpesviruses and the Unfolded Protein Response." Viruses 12, no. 1: 17.

Journal article
Published: 20 December 2019 in Viruses
Reads 0
Downloads 0

Kaposi’s sarcoma associated-herpesvirus (KSHV, also known as human herpesvirus-8) is a gammaherpesvirus that establishes life-long infection in human B lymphocytes. KSHV infection is typically asymptomatic, but immunosuppression can predispose KSHV-infected individuals to primary effusion lymphoma (PEL); a malignancy driven by aberrant proliferation of latently infected B lymphocytes, and supported by pro-inflammatory cytokines and angiogenic factors produced by cells that succumb to lytic viral replication. Here, we report the development of the first in vivo model for a virally induced lymphoma in zebrafish, whereby KSHV-infected PEL tumor cells engraft and proliferate in the yolk sac of zebrafish larvae. Using a PEL cell line engineered to produce the viral lytic switch protein RTA in the presence of doxycycline, we demonstrate drug-inducible reactivation from KSHV latency in vivo, which enabled real-time observation and evaluation of latent and lytic phases of KSHV infection. In addition, we developed a sensitive droplet digital PCR method to monitor latent and lytic viral gene expression and host cell gene expression in xenografts. The zebrafish yolk sac is not well vascularized, and by using fluorogenic assays, we confirmed that this site provides a hypoxic environment that may mimic the microenvironment of some human tumors. We found that PEL cell proliferation in xenografts was dependent on the host hypoxia-dependent translation initiation factor, eukaryotic initiation factor 4E2 (eIF4E2). This demonstrates that the zebrafish yolk sac is a functionally hypoxic environment, and xenografted cells must switch to dedicated hypoxic gene expression machinery to survive and proliferate. The establishment of the PEL xenograft model enables future studies that exploit the innate advantages of the zebrafish as a model for genetic and pharmacologic screens.

ACS Style

Eric S. Pringle; Jaime Wertman; Nicole Melong; Andrew J. Coombs; Andrew L. Young; David O’Leary; Chansey Veinotte; Carolyn-Ann Robinson; Michael N. Ha; Graham Dellaire; Todd E. Druley; Craig McCormick; Jason N. Berman. The Zebrafish Xenograft Platform—A Novel Tool for Modeling KSHV-Associated Diseases. Viruses 2019, 12, 12 .

AMA Style

Eric S. Pringle, Jaime Wertman, Nicole Melong, Andrew J. Coombs, Andrew L. Young, David O’Leary, Chansey Veinotte, Carolyn-Ann Robinson, Michael N. Ha, Graham Dellaire, Todd E. Druley, Craig McCormick, Jason N. Berman. The Zebrafish Xenograft Platform—A Novel Tool for Modeling KSHV-Associated Diseases. Viruses. 2019; 12 (1):12.

Chicago/Turabian Style

Eric S. Pringle; Jaime Wertman; Nicole Melong; Andrew J. Coombs; Andrew L. Young; David O’Leary; Chansey Veinotte; Carolyn-Ann Robinson; Michael N. Ha; Graham Dellaire; Todd E. Druley; Craig McCormick; Jason N. Berman. 2019. "The Zebrafish Xenograft Platform—A Novel Tool for Modeling KSHV-Associated Diseases." Viruses 12, no. 1: 12.

Journal article
Published: 17 December 2019 in Infection and Immunity
Reads 0
Downloads 0

Pseudomonas aeruginosa is an opportunistic pathogen that is a common cause of nosocomial infections. The molecular mechanisms governing immune responses to P. aeruginosa infection remain incompletely defined. Early growth response 1 (Egr-1) is a zinc-finger transcription factor that controls inflammatory responses. Here, we characterized the role of Egr-1 in host defense against P. aeruginosa infection in a mouse model of acute bacterial pneumonia.

ACS Style

Zheng Pang; Renee Raudonis; Craig McCormick; Zhenyu Cheng. Early Growth Response 1 Deficiency Protects the Host against Pseudomonas aeruginosa Lung Infection. Infection and Immunity 2019, 88, 1 .

AMA Style

Zheng Pang, Renee Raudonis, Craig McCormick, Zhenyu Cheng. Early Growth Response 1 Deficiency Protects the Host against Pseudomonas aeruginosa Lung Infection. Infection and Immunity. 2019; 88 (1):1.

Chicago/Turabian Style

Zheng Pang; Renee Raudonis; Craig McCormick; Zhenyu Cheng. 2019. "Early Growth Response 1 Deficiency Protects the Host against Pseudomonas aeruginosa Lung Infection." Infection and Immunity 88, no. 1: 1.

Research article
Published: 02 December 2019 in PLOS Pathogens
Reads 0
Downloads 0

Herpesviruses usurp host cell protein synthesis machinery to convert viral mRNAs into proteins, and the endoplasmic reticulum (ER) to ensure proper folding, post-translational modification and trafficking of secreted and transmembrane viral proteins. Overloading ER folding capacity activates the unfolded protein response (UPR), whereby sensor proteins ATF6, PERK and IRE1 initiate a stress-mitigating transcription program that accelerates catabolism of misfolded proteins while increasing ER folding capacity. Kaposi’s sarcoma-associated herpesvirus (KSHV) can be reactivated from latency by chemical induction of ER stress, which causes accumulation of the XBP1s transcription factor that transactivates the viral RTA lytic switch gene. The presence of XBP1s-responsive elements in the RTA promoter suggests that KSHV evolved a mechanism to respond to ER stress. Here, we report that ATF6, PERK and IRE1 were activated upon reactivation from latency and required for efficient KSHV lytic replication; genetic or pharmacologic inhibition of each UPR sensor diminished virion production. Despite UPR sensor activation during KSHV lytic replication, downstream UPR transcriptional responses were restricted; 1) ATF6 was cleaved to activate the ATF6(N) transcription factor but ATF6(N)-responsive genes were not transcribed; 2) PERK phosphorylated eIF2α but ATF4 did not accumulate; 3) IRE1 caused XBP1 mRNA splicing, but XBP1s protein did not accumulate and XBP1s-responsive genes were not transcribed. Ectopic expression of the KSHV host shutoff protein SOX did not affect UPR gene expression, suggesting that alternative viral mechanisms likely mediate UPR suppression during lytic replication. Complementation of XBP1s deficiency during KSHV lytic replication inhibited virion production in a dose-dependent manner in iSLK.219 cells but not in TREx-BCBL1-RTA cells. However, genetically distinct KSHV virions harvested from these two cell lines were equally susceptible to XBP1s restriction following infection of naïve iSLK cells. This suggests that cell-intrinsic properties of BCBL1 cells may circumvent the antiviral effect of ectopic XBP1s expression. Taken together, these findings indicate that while XBP1s plays an important role in reactivation from latency, it can inhibit virus replication at a later step, which the virus overcomes by preventing its synthesis. These findings suggest that KSHV hijacks UPR sensors to promote efficient viral replication while sustaining ER stress. Like all viruses, Kaposi’s sarcoma-associated herpesvirus (KSHV) uses cellular machinery to create viral proteins. Some of these proteins are folded and modified in the endoplasmic reticulum (ER) and traverse the cellular secretory apparatus. Exceeding ER protein folding capacity activates the unfolded protein response (UPR), which resolves ER stress by putting the brakes on protein synthesis and turning on stress-mitigating genes. We show that KSHV replication activates the three cellular proteins that sense ER stress, which are each required to support efficient viral replication. By contrast, KSHV blocks the UPR gene expression program downstream from each of these activated sensor proteins. The failure to resolve ER stress might normally be expected to put the virus at a disadvantage, but we demonstrate that reversal of this scenario is worse; when we supplement infected epithelial cells with the UPR transcription factor XBP1s to artificially stimulate the production of UPR-responsive gene products, virus replication is blocked at a late stage and very few viruses are released from infected cells. Taken together, these observations suggest that KSHV requires UPR sensor protein activation to replicate but has dramatically altered the outcome to prevent the synthesis of new UPR proteins and sustain stress in the ER compartment.

ACS Style

Benjamin P. Johnston; Eric S. Pringle; Craig McCormick. KSHV activates unfolded protein response sensors but suppresses downstream transcriptional responses to support lytic replication. PLOS Pathogens 2019, 15, e1008185 .

AMA Style

Benjamin P. Johnston, Eric S. Pringle, Craig McCormick. KSHV activates unfolded protein response sensors but suppresses downstream transcriptional responses to support lytic replication. PLOS Pathogens. 2019; 15 (12):e1008185.

Chicago/Turabian Style

Benjamin P. Johnston; Eric S. Pringle; Craig McCormick. 2019. "KSHV activates unfolded protein response sensors but suppresses downstream transcriptional responses to support lytic replication." PLOS Pathogens 15, no. 12: e1008185.

Journal article
Published: 01 November 2019 in Journal of Virology
Reads 0
Downloads 0

All viruses require host cell machinery to synthesize viral proteins. A host cell protein complex known as mechanistic target of rapamycin complex 1 (mTORC1) is a master regulator of protein synthesis. Under nutrient-rich conditions, mTORC1 is active and promotes protein synthesis to meet cellular anabolic demands. Under nutrient-poor conditions or under stress, mTORC1 is rapidly inhibited, global protein synthesis is arrested, and a cellular catabolic process known as autophagy is activated. Kaposi’s sarcoma-associated herpesvirus (KSHV) stimulates mTORC1 activity and utilizes host machinery to synthesize viral proteins. However, we discovered that mTORC1 activity was largely dispensable for viral protein synthesis, genome replication, and the release of infectious progeny. Likewise, during lytic replication, mTORC1 was no longer able to control autophagy. These findings suggest that KSHV undermines mTORC1-dependent cellular processes during the lytic cycle to ensure efficient viral replication.

ACS Style

Eric S. Pringle; Carolyn-Ann Robinson; Craig McCormick. Kaposi’s Sarcoma-Associated Herpesvirus Lytic Replication Interferes with mTORC1 Regulation of Autophagy and Viral Protein Synthesis. Journal of Virology 2019, 93, 1 .

AMA Style

Eric S. Pringle, Carolyn-Ann Robinson, Craig McCormick. Kaposi’s Sarcoma-Associated Herpesvirus Lytic Replication Interferes with mTORC1 Regulation of Autophagy and Viral Protein Synthesis. Journal of Virology. 2019; 93 (21):1.

Chicago/Turabian Style

Eric S. Pringle; Carolyn-Ann Robinson; Craig McCormick. 2019. "Kaposi’s Sarcoma-Associated Herpesvirus Lytic Replication Interferes with mTORC1 Regulation of Autophagy and Viral Protein Synthesis." Journal of Virology 93, no. 21: 1.

Article
Published: 01 April 2019 in Cell Reports
Reads 0
Downloads 0

Summary Many viruses shut off host gene expression to inhibit antiviral responses. Viral proteins and host proteins required for viral replication are typically spared in this process, but the mechanisms of target selectivity during host shutoff remain poorly understood. Using transcriptome-wide and targeted reporter experiments, we demonstrate that the influenza A virus endoribonuclease PA-X usurps RNA splicing to selectively target host RNAs for destruction. Proximity-labeling proteomics reveals that PA-X interacts with cellular RNA processing proteins, some of which are partially required for host shutoff. Thus, PA-X taps into host nuclear pre-mRNA processing mechanisms to destroy nascent mRNAs shortly after their synthesis. This mechanism sets PA-X apart from other viral host shutoff proteins that target actively translating mRNAs in the cytoplasm. Our study reveals a unique mechanism of host shutoff that helps us understand how influenza viruses suppress host gene expression.

ACS Style

Lea Gaucherand; Brittany K. Porter; Rachel E. Levene; Emma L. Price; Summer K. Schmaling; Chris H. Rycroft; Yuzo Kevorkian; Craig McCormick; Denys A. Khaperskyy; Marta M. Gaglia. The Influenza A Virus Endoribonuclease PA-X Usurps Host mRNA Processing Machinery to Limit Host Gene Expression. Cell Reports 2019, 27, 776 -792.e7.

AMA Style

Lea Gaucherand, Brittany K. Porter, Rachel E. Levene, Emma L. Price, Summer K. Schmaling, Chris H. Rycroft, Yuzo Kevorkian, Craig McCormick, Denys A. Khaperskyy, Marta M. Gaglia. The Influenza A Virus Endoribonuclease PA-X Usurps Host mRNA Processing Machinery to Limit Host Gene Expression. Cell Reports. 2019; 27 (3):776-792.e7.

Chicago/Turabian Style

Lea Gaucherand; Brittany K. Porter; Rachel E. Levene; Emma L. Price; Summer K. Schmaling; Chris H. Rycroft; Yuzo Kevorkian; Craig McCormick; Denys A. Khaperskyy; Marta M. Gaglia. 2019. "The Influenza A Virus Endoribonuclease PA-X Usurps Host mRNA Processing Machinery to Limit Host Gene Expression." Cell Reports 27, no. 3: 776-792.e7.

Meeting report
Published: 18 January 2019 in Viruses
Reads 0
Downloads 0

The 2nd Symposium of the Canadian Society for Virology (CSV2018) was held in June 2018 in Halifax, Nova Scotia, Canada, as a featured event marking the 200th anniversary of Dalhousie University. CSV2018 attracted 175 attendees from across Canada and around the world, more than double the number that attended the first CSV symposium two years earlier. CSV2018 provided a forum to discuss a wide range of topics in virology including human, veterinary, plant, and microbial pathogens. Invited keynote speakers included David Kelvin (Dalhousie University and Shantou University Medical College) who provided a historical perspective on influenza on the 100th anniversary of the 1918 pandemic; Sylvain Moineau (Université Laval) who described CRISPR-Cas systems and anti-CRISPR proteins in warfare between bacteriophages and their host microbes; and Kate O’Brien (then from Johns Hopkins University, now relocated to the World Health Organization where she is Director of Immunization, Vaccines and Biologicals), who discussed the underlying viral etiology for pneumonia in the developing world, and the evidence for respiratory syncytial virus (RSV) as a primary cause. Reflecting a strong commitment of Canadian virologists to science communication, CSV2018 featured the launch of Halifax’s first annual Soapbox Science event to enable public engagement with female scientists, and the live-taping of the 499th episode of the This Week in Virology (TWIV) podcast, hosted by Vincent Racaniello (Columbia University) and science writer Alan Dove. TWIV featured interviews of CSV co-founders Nathalie Grandvaux (Université de Montréal) and Craig McCormick (Dalhousie University), who discussed the origins and objectives of the new society; Ryan Noyce (University of Alberta), who discussed technical and ethical considerations of synthetic virology; and Kate O’Brien, who discussed vaccines and global health. Finally, because CSV seeks to provide a better future for the next generation of Canadian virologists, the symposium featured a large number of oral and poster presentations from trainees and closed with the awarding of presentation prizes to trainees, followed by a tour of the Halifax Citadel National Historic Site and an evening of entertainment at the historic Alexander Keith’s Brewery.

ACS Style

Nathalie Grandvaux; Craig McCormick. CSV2018: The 2nd Symposium of the Canadian Society for Virology. Viruses 2019, 11, 79 .

AMA Style

Nathalie Grandvaux, Craig McCormick. CSV2018: The 2nd Symposium of the Canadian Society for Virology. Viruses. 2019; 11 (1):79.

Chicago/Turabian Style

Nathalie Grandvaux; Craig McCormick. 2019. "CSV2018: The 2nd Symposium of the Canadian Society for Virology." Viruses 11, no. 1: 79.

Research article
Published: 02 January 2019 in PLOS ONE
Reads 0
Downloads 0

Plant cell walls are composed of cellulose, hemicellulose, and lignin, collectively known as lignocellulose. Microorganisms degrade lignocellulose to liberate sugars to meet metabolic demands. Using a metagenomic sequencing approach, we previously demonstrated that the microbiome of the North American porcupine (Erethizon dorsatum) is replete with genes that could encode lignocellulose-degrading enzymes. Here, we report the identification, synthesis and partial characterization of four novel genes from the porcupine microbiome encoding putative lignocellulose-degrading enzymes: β-glucosidase, α-L-arabinofuranosidase, β-xylosidase, and endo-1,4-β-xylanase. These genes were identified via conserved catalytic domains associated with cellulose- and hemicellulose-degradation. Phylogenetic trees were created for each of these putative enzymes to depict genetic relatedness to known enzymes. Candidate genes were synthesized and cloned into plasmid expression vectors for inducible protein expression and secretion. The putative β-glucosidase fusion protein was efficiently secreted but did not permit Escherichia coli (E. coli) to use cellobiose as a sole carbon source, nor did the affinity purified enzyme cleave p-Nitrophenyl β-D-glucopyranoside (p-NPG) substrate in vitro over a range of physiological pH levels (pH 5–7). The putative hemicellulose-degrading β-xylosidase and α-L-arabinofuranosidase enzymes also lacked in vitro enzyme activity, but the affinity purified endo-1,4-β-xylanase protein cleaved a 6-chloro-4-methylumbelliferyl xylobioside substrate in acidic and neutral conditions, with maximal activity at pH 7. At this optimal pH, KM, Vmax, and kcat were determined to be 32.005 ± 4.72 μM, 1.16x10-5 ± 3.55x10-7 M/s, and 94.72 s-1, respectively. Thus, our pipeline enabled successful identification and characterization of a novel hemicellulose-degrading enzyme from the porcupine microbiome. Progress towards the goal of introducing a complete lignocellulose-degradation pathway into E. coli will be accelerated by combining synthetic metagenomic approaches with functional metagenomic library screening, which can identify novel enzymes unrelated to those found in available databases.

ACS Style

Mackenzie Thornbury; Jacob Sicheri; Patrick Slaine; Landon J. Getz; Emma Finlayson-Trick; Jamie Cook; Caroline Guinard; Nicholas Boudreau; David Jakeman; John Rohde; Craig McCormick. Characterization of novel lignocellulose-degrading enzymes from the porcupine microbiome using synthetic metagenomics. PLOS ONE 2019, 14, e0209221 .

AMA Style

Mackenzie Thornbury, Jacob Sicheri, Patrick Slaine, Landon J. Getz, Emma Finlayson-Trick, Jamie Cook, Caroline Guinard, Nicholas Boudreau, David Jakeman, John Rohde, Craig McCormick. Characterization of novel lignocellulose-degrading enzymes from the porcupine microbiome using synthetic metagenomics. PLOS ONE. 2019; 14 (1):e0209221.

Chicago/Turabian Style

Mackenzie Thornbury; Jacob Sicheri; Patrick Slaine; Landon J. Getz; Emma Finlayson-Trick; Jamie Cook; Caroline Guinard; Nicholas Boudreau; David Jakeman; John Rohde; Craig McCormick. 2019. "Characterization of novel lignocellulose-degrading enzymes from the porcupine microbiome using synthetic metagenomics." PLOS ONE 14, no. 1: e0209221.

Unit
Published: 29 October 2018 in Current Protocols in Molecular Biology
Reads 0
Downloads 0

Post‐transcriptional regulation is an important aspect of the control of gene expression. mRNAs are translated with variable efficiencies, and these efficiencies can change rapidly during adaptation to diverse environmental factors, including cellular stresses and microbial infections. Polysome profiling analysis utilizes ultracentrifugation to isolate complexes of mRNAs in the process of translation and corresponding proteins on the basis of density. Here we describe polysome profiling analysis using a continuous ultraviolet spectrophotometer and a gradient fractionator. We provide protocols for processing sucrose gradient fractions for isolation of RNA for RT‐qPCR analysis and isolation of protein for SDS‐PAGE and immunoblot analysis. © 2018 by John Wiley & Sons, Inc.

ACS Style

Eric S. Pringle; Craig McCormick; Zhenyu Cheng. Polysome Profiling Analysis of mRNA and Associated Proteins Engaged in Translation. Current Protocols in Molecular Biology 2018, 125, e79 .

AMA Style

Eric S. Pringle, Craig McCormick, Zhenyu Cheng. Polysome Profiling Analysis of mRNA and Associated Proteins Engaged in Translation. Current Protocols in Molecular Biology. 2018; 125 (1):e79.

Chicago/Turabian Style

Eric S. Pringle; Craig McCormick; Zhenyu Cheng. 2018. "Polysome Profiling Analysis of mRNA and Associated Proteins Engaged in Translation." Current Protocols in Molecular Biology 125, no. 1: e79.

Other
Published: 14 October 2018
Reads 0
Downloads 0

SUMMARYMany viruses globally shut off host gene expression to inhibit activation of cell-intrinsic antiviral responses. However, host shutoff is not indiscriminate, since viral proteins and host proteins required for viral replication are still synthesized during shutoff. The molecular determinants of target selectivity in host shutoff remain incompletely understood. Here, we report that the influenza A virus shutoff factor PA-X usurps RNA splicing to selectively target host RNAs for destruction. PA-X preferentially degrades spliced mRNAs, both transcriptome-wide and in reporter assays. Moreover, proximity-labeling proteomics revealed that PA-X interacts with cellular proteins involved in RNA splicing. The interaction with splicing contributes to target discrimination and is unique among viral host shutoff nucleases. This novel mechanism sheds light on the specificity of viral control of host gene expression and may provide opportunities for development of new host-targeted antivirals.

ACS Style

Lea Gaucherand; Brittany K. Porter; Summer K. Schmaling; Christopher Harley Rycroft; Yuzo Kevorkian; Craig McCormick; Denys A. Khaperskyy; Marta Maria Gaglia. The influenza A virus endoribonuclease PA-X usurps host mRNA processing machinery to limit host gene expression. 2018, 442996 .

AMA Style

Lea Gaucherand, Brittany K. Porter, Summer K. Schmaling, Christopher Harley Rycroft, Yuzo Kevorkian, Craig McCormick, Denys A. Khaperskyy, Marta Maria Gaglia. The influenza A virus endoribonuclease PA-X usurps host mRNA processing machinery to limit host gene expression. . 2018; ():442996.

Chicago/Turabian Style

Lea Gaucherand; Brittany K. Porter; Summer K. Schmaling; Christopher Harley Rycroft; Yuzo Kevorkian; Craig McCormick; Denys A. Khaperskyy; Marta Maria Gaglia. 2018. "The influenza A virus endoribonuclease PA-X usurps host mRNA processing machinery to limit host gene expression." , no. : 442996.

Preprint
Published: 12 October 2018
Reads 0
Downloads 0

Herpesviruses usurp host cell protein synthesis machinery to convert viral mRNAs into proteins, and the endoplasmic reticulum (ER) to ensure proper folding, post-translational modification and trafficking of secreted viral proteins. Overloading ER folding capacity activates the unfolded protein response (UPR), whereby displacement of the ER chaperone BiP activates UPR sensor proteins ATF6, PERK and IRE1 to initiate transcriptional responses to increase catabolic processes and ER folding capacity, while suppressing bulk protein synthesis. Kaposi’s sarcoma-associated herpesvirus (KSHV) can be reactivated from latency by chemical induction of ER stress, whereby the IRE1 endoribonuclease cleaves XBP1 mRNA, resulting in a ribosomal frameshift that yields the XBP1s transcription factor that transactivates the promoter of K-RTA, the viral lytic switch protein. By incorporating XBP1s responsive elements in the K-RTA promoter KSHV appears to have evolved a mechanism to respond to ER stress. Here, we report that following reactivation from latency, KSHV lytic replication causes activation of ATF6, PERK and IRE1 UPR sensor proteins. UPR sensor activation is required for efficient KSHV lytic replication; genetic or pharmacologic inhibition of each UPR sensor diminishes virion production. Despite strong UPR sensor activation during KSHV lytic replication, downstream UPR transcriptional responses were restricted; 1) ATF6 was cleaved to release the ATF6(N) transcription factor but known ATF6(N)-responsive genes were not transcribed; 2) PERK phosphorylated eIF2α but ATF4 did not accumulate as expected; 3) IRE1 caused XBP1 mRNA splicing, but XBP1s protein failed to accumulate and XBP1s-responsive genes were not transcribed. Remarkably, complementation of XBP1s deficiency during KSHV lytic replication by ectopic expression inhibited the production of infectious virions in a dose-dependent manner. Therefore, while XBP1s plays an important role in reactivation from latency, it inhibits later steps in lytic replication, which the virus overcomes by preventing its synthesis. Taken together, these findings suggest that KSHV hijacks UPR sensors to promote efficient viral replication while sustaining ER stress.Author summaryHuman herpesvirus-8 is the most recently discovered human herpesvirus, and it is the infectious cause of Kaposi’s sarcoma, which is why it’s also known as Kaposi’s sarcoma-associated herpesvirus (KSHV). Like all herpesviruses, KSHV replicates in the cell nucleus and uses host cell machinery to convert viral genes into proteins. Some of these proteins are synthesized, folded and modified in the endoplasmic reticulum (ER) and traverse the cellular secretory apparatus. Because the virus heavily utilizes the ER to make and process proteins, there is potential to overwhelm the system, which could impede viral replication and in extreme cases, kill the cell. Normally, when demands on the protein folding machinery are exceeded then misfolded...

ACS Style

Benjamin P. Johnston; Craig McCormick. KSHV activates unfolded protein response sensors but suppresses downstream transcriptional responses to support lytic replication. 2018, 442079 .

AMA Style

Benjamin P. Johnston, Craig McCormick. KSHV activates unfolded protein response sensors but suppresses downstream transcriptional responses to support lytic replication. . 2018; ():442079.

Chicago/Turabian Style

Benjamin P. Johnston; Craig McCormick. 2018. "KSHV activates unfolded protein response sensors but suppresses downstream transcriptional responses to support lytic replication." , no. : 442079.