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Li Y, Zhou X, Qi S, Jia G, Cao J, Guan Z, Zhao R. Effects of Portulaca oleracea L. Polysaccharide on piglets infected with porcine rotavirus. Microb Pathog 2025; 200:107355. [PMID: 39892834 DOI: 10.1016/j.micpath.2025.107355] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Academic Contribution Register] [Received: 09/25/2024] [Revised: 01/05/2025] [Accepted: 01/30/2025] [Indexed: 02/04/2025]
Abstract
Piglet diarrhea stands as the primary cause of piglet mortality, inflicting substantial economic losses to pig farmers. Porcine rotavirus (PoRV), a member of the Reoviridae family, bears responsibility for instigating diarrhea and severe dehydration, culminating in piglet fatalities. Portulaca oleracea L. (POL), a Chinese medicinal herb commonly utilized for its antiviral properties and effectiveness in treating diarrhea, holds promise for mitigating these effects. Nevertheless, the in vivo antiviral effect of Portulaca oleracea L. polysaccharide (POL-P), a key component of POL, on PoRV infection remains obscure. This study aimed to investigate the antiviral effects of POL-P on PoRV in piglets and its impact on their intestinal flora. Studies have demonstrated that oral POL-P improves growth in both lactating mice and PoRV-infected piglets, while reducing diarrhea and mortality. It also helps to reduce intestinal damage and lower viral mRNA levels in the jejunum and ileum. Additionally, POL-P improves blood tests in piglets, boosting levels of IFN-α and IL-10, and lowering tumor necrosis factor-α and IL-6. Analysis of the intestinal flora in PoRV-infected piglets showed reduced levels of Bacteroides (18.39 %) and increased levels of Firmicutes (39.81 %), Proteobacteria (33.46 %), and Verrucomicrobia (7.42 %). After POL-P treatment, Bacteroides increased to 28.31 %, while Firmicutes, Proteobacteria, and Verrucomicrobia decreased to 34.93 %, 25.52 %, and 2.53 %, respectively. At the genus level, POL-P treatment reduced the abundance of Klebsiella, Ackermannia, and Streptococcus. In conclusion, POL-P helps reduce inflammation and intestinal damage caused by PoRV infection, prevents viral colonization, restores gut flora balance, improves piglet growth, and reduces diarrhea and mortality.
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Affiliation(s)
- Yan Li
- College of Life Science & Biotechnology, Heilongjiang Bayi Agricultural University, Daqing High-Tech Industrial Development Zone, 163319, PR China
| | - Xiechen Zhou
- College of Animal Science and Technology, Heilongjiang Bayi Agricultural University, Daqing High-Tech Industrial Development Zone, 163319, PR China
| | - Shanshan Qi
- College of Animal Science and Technology, Heilongjiang Bayi Agricultural University, Daqing High-Tech Industrial Development Zone, 163319, PR China
| | - Guiyan Jia
- College of Life Science & Biotechnology, Heilongjiang Bayi Agricultural University, Daqing High-Tech Industrial Development Zone, 163319, PR China
| | - Junyang Cao
- College of Life Science & Biotechnology, Heilongjiang Bayi Agricultural University, Daqing High-Tech Industrial Development Zone, 163319, PR China
| | - Zijan Guan
- College of Life Science & Biotechnology, Heilongjiang Bayi Agricultural University, Daqing High-Tech Industrial Development Zone, 163319, PR China
| | - Rui Zhao
- College of Life Science & Biotechnology, Heilongjiang Bayi Agricultural University, Daqing High-Tech Industrial Development Zone, 163319, PR China; Heilongjiang Provincial Key Laboratory of Animal Cell Activities and Stress Adaptations, Heilongjiang Bayi Agricultural University, Daqing High-Tech Industrial Development Zone, 163319, PR China.
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Jung J, Bae J, Park JS, Lee SW, Jeong JH, Park SJ. In Vitro Anti-Rotaviral Activity of Bavachin Isolated from Psoralea corylifolia L. ( Fabaceae). Vet Sci 2024; 11:188. [PMID: 38787160 PMCID: PMC11126063 DOI: 10.3390/vetsci11050188] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Academic Contribution Register] [Received: 03/21/2024] [Revised: 04/12/2024] [Accepted: 04/24/2024] [Indexed: 05/25/2024] Open
Abstract
Rotavirus is the main causative agent of viral gastroenteritis among young animals worldwide. Currently, no clinically approved or effective antiviral drugs are available to combat rotavirus infections. Herein, we evaluated the anti-rotaviral activities of extracts and bavachin isolated from Psoralea corylifolia L. (Fabaceae) (P. corylifolia) against the bovine rotavirus G8P[7] and porcine rotavirus G5P[7] in vitro. Two assay strategies were performed: (1) a virucidal assay to reduce viral infectivity by virus neutralization and (2) a post-treatment assay to assess viral replication suppression. The results from the virucidal assay showed that the extracts and bavachin did not exert anti-rotaviral activities. In the follow-up analysis after treatment, bavachin exhibited robust antiviral efficacy, with 50% effective concentration (EC50) values of 10.6 μM (selectivity index [SI] = 2.38) against bovine rotavirus G8P[7] and 13.0 μM (SI = 1.94) against porcine rotavirus G5P[7]. Bavachin strongly suppressed viral RNA synthesis in the early (6 h) and late stages (18 h) after rotaviral infection. These findings strongly suggest that bavachin may have hindered the virions by effectively inhibiting the early stages of the virus replication cycle after rotaviral infection. Furthermore, confocal imaging showed that bavachin suppressed viral protein synthesis, notably that of the rotaviral protein (VP6). These results suggest that bavachin has strong antiviral activity against rotaviruses, inhibits viral replication, and is a candidate natural therapeutic drug targeting rotaviral infection. The utilization of bavachin isolated from P. corylifolia may contribute to decreased mortality rates, lower medication expenses, and enhanced economic viability in domestic farms.
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Affiliation(s)
- Jinseok Jung
- Functional Biomaterial Research Center, Korea Research Institute of Bioscience and Biotechnology, 181 Ipsin-gil, Jeongeup-si 56212, Republic of Korea; (J.J.); (J.B.); (J.S.P.); (S.W.L.)
- Department of Microbiology, Chonnam National University Medical School, Hwasun 58128, Republic of Korea
| | - Jaehoon Bae
- Functional Biomaterial Research Center, Korea Research Institute of Bioscience and Biotechnology, 181 Ipsin-gil, Jeongeup-si 56212, Republic of Korea; (J.J.); (J.B.); (J.S.P.); (S.W.L.)
- Department of Microbiology, Chonnam National University Medical School, Hwasun 58128, Republic of Korea
| | - Ji Sun Park
- Functional Biomaterial Research Center, Korea Research Institute of Bioscience and Biotechnology, 181 Ipsin-gil, Jeongeup-si 56212, Republic of Korea; (J.J.); (J.B.); (J.S.P.); (S.W.L.)
| | - Seung Woong Lee
- Functional Biomaterial Research Center, Korea Research Institute of Bioscience and Biotechnology, 181 Ipsin-gil, Jeongeup-si 56212, Republic of Korea; (J.J.); (J.B.); (J.S.P.); (S.W.L.)
| | - Jae-Ho Jeong
- Department of Microbiology, Chonnam National University Medical School, Hwasun 58128, Republic of Korea
| | - Su-Jin Park
- Functional Biomaterial Research Center, Korea Research Institute of Bioscience and Biotechnology, 181 Ipsin-gil, Jeongeup-si 56212, Republic of Korea; (J.J.); (J.B.); (J.S.P.); (S.W.L.)
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Raev S, Amimo J, Saif L, Vlasova A. Intestinal mucin-type O-glycans: the major players in the host-bacteria-rotavirus interactions. Gut Microbes 2023; 15:2197833. [PMID: 37020288 PMCID: PMC10078158 DOI: 10.1080/19490976.2023.2197833] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Academic Contribution Register] [Received: 12/07/2022] [Accepted: 03/28/2023] [Indexed: 04/07/2023] Open
Abstract
Rotavirus (RV) causes severe diarrhea in young children and animals worldwide. Several glycans terminating in sialic acids (SAs) and histo-blood group antigens (HBGAs) on intestinal epithelial cell (IEC) surface have been recognized to act as attachment sites for RV. IECs are protected by the double layer of mucus of which O-glycans (including HBGAs and SAs) are a major organic component. Luminal mucins, as well as bacterial glycans, can act as decoy molecules removing RV particles from the gut. The composition of the intestinal mucus is regulated by complex O-glycan-specific interactions among the gut microbiota, RV and the host. In this review, we highlight O-glycan-mediated interactions within the intestinal lumen prior to RV attachment to IECs. A better understanding of the role of mucus is essential for the development of alternative therapeutic tools including the use of pre- and probiotics to control RV infection.
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Affiliation(s)
- S.A. Raev
- Center for Food Animal Health, Department of Animal Sciences, College of Food, Agricultural and Environmental Sciences, The Ohio State University, Wooster, OH, USA
| | - J.O. Amimo
- Center for Food Animal Health, Department of Animal Sciences, College of Food, Agricultural and Environmental Sciences, The Ohio State University, Wooster, OH, USA
- Department of Animal Production, Faculty of Veterinary Medicine, University of Nairobi, Nairobi, Kenya
| | - L.J. Saif
- Center for Food Animal Health, Department of Animal Sciences, College of Food, Agricultural and Environmental Sciences, The Ohio State University, Wooster, OH, USA
| | - A.N. Vlasova
- Center for Food Animal Health, Department of Animal Sciences, College of Food, Agricultural and Environmental Sciences, The Ohio State University, Wooster, OH, USA
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Patra U, Mukhopadhyay U, Mukherjee A, Dutta S, Chawla-Sarkar M. Treading a HOSTile path: Mapping the dynamic landscape of host cell-rotavirus interactions to explore novel host-directed curative dimensions. Virulence 2021; 12:1022-1062. [PMID: 33818275 PMCID: PMC8023246 DOI: 10.1080/21505594.2021.1903198] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Academic Contribution Register] [Received: 09/21/2020] [Revised: 01/20/2021] [Accepted: 03/10/2021] [Indexed: 12/27/2022] Open
Abstract
Viruses are intracellular pathogens and are dependent on host cellular resources to carry out their cycles of perpetuation. Obtaining an integrative view of host-virus interaction is of utmost importance to understand the complex and dynamic interplay between viral components and host machineries. Besides its obvious scholarly significance, a comprehensive host-virus interaction profile also provides a platform where from host determinants of pro-viral and antiviral importance can be identified and further be subjected to therapeutic intervention. Therefore, adjunct to conventional methods of prophylactic vaccination and virus-directed antivirals, this host-targeted antiviral approach holds promising therapeutic potential. In this review, we present a comprehensive landscape of host cellular reprogramming in response to infection with rotavirus (RV) which causes profuse watery diarrhea in neonates and infants. In addition, an emphasis is given on how host determinants are either usurped or subverted by RV in course of infection and how therapeutic manipulation of specific host factors can effectively modulate the RV life cycle.
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Affiliation(s)
- Upayan Patra
- Division of Virology, National Institute of Cholera and Enteric Diseases, Beliaghata, Kolkata, India
| | - Urbi Mukhopadhyay
- Division of Virology, National Institute of Cholera and Enteric Diseases, Beliaghata, Kolkata, India
| | - Arpita Mukherjee
- Division of Virology, National Institute of Cholera and Enteric Diseases, Beliaghata, Kolkata, India
| | - Shanta Dutta
- Division of Bacteriology, National Institute of Cholera and Enteric Diseases, Beliaghata, Kolkata, India
| | - Mamta Chawla-Sarkar
- Division of Virology, National Institute of Cholera and Enteric Diseases, Beliaghata, Kolkata, India
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Abstract
Group A rotavirus (RVA), one of the leading pathogens causing severe acute gastroenteritis in children and a wide variety of young animals worldwide, induces apoptosis upon infecting cells. Though RVA-induced apoptosis mediated via the dual modulation of its NSP4 and NSP1 proteins is relatively well studied, the nature and signaling pathway(s) involved in RVA-induced necroptosis are yet to be fully elucidated. Here, we demonstrate the nature of RVA-induced necroptosis, the signaling cascade involved, and correlation with RVA-induced apoptosis. Infection with the bovine NCDV and human DS-1 RV strains was shown to activate receptor-interacting protein kinase 1 (RIPK1)/RIPK3/mixed lineage kinase domain-like protein (MLKL), the key necroptosis molecules in virus-infected cells. Using immunoprecipitation assay, RIPK1 was found to bind phosphorylated RIPK3 (pRIPK3) and pMLKL. pMLKL, the major executioner molecule in the necroptotic pathway, was translocated to the plasma membrane of RVA-infected cells to puncture the cell membrane. Interestingly, transfection of RVA NSP4 also induced necroptosis through the RIPK1/RIPK3/MLKL necroptosis pathway. Blockage of each key necroptosis molecule in the RVA-infected or NSP4-transfected cells resulted in decreased necroptosis but increased cell viability and apoptosis, thereby resulting in decreased viral yields in the RVA-infected cells. In contrast, suppression of RVA-induced apoptosis increased necroptosis and virus yields. Our findings suggest that RVA NSP4 also induces necroptosis via the RIPK1/RIPK3/MLKL necroptosis pathway. Moreover, necroptosis and apoptosis-which have proviral and antiviral effects, respectively-exhibited a crosstalk in RVA-infected cells. These findings significantly increase our understanding of the nature of RVA-induced necroptosis and the crosstalk between RVA-induced necroptosis and apoptosis. IMPORTANCE Viral infection usually culminates in cell death through apoptosis, necroptosis, and rarely, pyroptosis. Necroptosis is a form of programmed necrosis that is mediated by signaling complexes of the receptor-interacting protein kinase 1 (RIPK1), RIPK3, and mixed lineage kinase domain-like protein (MLKL). Although apoptosis induction by rotavirus and its NSP4 protein is well known, rotavirus-induced necroptosis is not fully understood. Here, we demonstrate that rotavirus and also its NSP4 protein can induce necroptosis in cultured cells through the activation of the RIPK1/RIPK3/MLKL necroptosis pathway. Moreover, rotavirus-induced necroptosis and apoptosis have opposite effects on viral yield, i.e., they function as proviral and antiviral processes, respectively, and counterbalance each other in rotavirus-infected cells. Our findings provide important insights for understanding the nature of rotavirus-induced necroptosis and the development of novel therapeutic strategies against infection with rotavirus and other RNA viruses.
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Progressive Rotavirus Infection Downregulates Redox-Sensitive Transcription Factor Nrf2 and Nrf2-Driven Transcription Units. OXIDATIVE MEDICINE AND CELLULAR LONGEVITY 2020; 2020:7289120. [PMID: 32322337 PMCID: PMC7165344 DOI: 10.1155/2020/7289120] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Academic Contribution Register] [Received: 10/16/2019] [Revised: 01/31/2020] [Accepted: 02/10/2020] [Indexed: 12/13/2022]
Abstract
Eukaryotic cells adopt highly tuned stress response physiology under threats of exogenous stressors including viruses to maintain cellular homeostasis. Not surprisingly, avoidance of cellular stress response pathways is an essential facet of virus-induced obligatory host reprogramming to invoke a cellular environment conducive to viral perpetuation. Adaptive cellular responses to oxidative and electrophilic stress are usually taken care of by an antioxidant defense system, core to which lies the redox-responsive transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2) and Nrf2-driven transcriptional cascade. Deregulation of host redox balance and redox stress-sensitive Nrf2 antioxidant defense have been reported for many viruses. In the current study, we aimed to study the modulation of the Nrf2-based host cellular redox defense system in response to Rotavirus (RV) infection in vitro. Interestingly, we found that Nrf2 protein levels decline sharply with progression of RV infection beyond an initial upsurge. Moreover, Nrf2 decrease as a whole was found to be accompanied by active nuclear vacuity of Nrf2, resulting in lowered expression of stress-responsive Nrf2 target genes heme oxygenase-1 (HO-1), NAD(P)H quinone dehydrogenase 1, and superoxide dismutase 1 both in the presence and absence of Nrf2-driven transcriptional inducers. Initial induction of Nrf2 concurred with RV-induced early burst of oxidative stress and therefore was sensitive to treatments with antioxidants. Reduction of Nrf2 levels beyond initial hours, however, was found to be independent of the cellular redox status. Furthermore, increasing the half-life of Nrf2 through inhibition of the Kelch-like erythroid cell-derived protein with CNC homology- (ECH-) associated protein 1/Cullin3-RING Box1-based canonical Nrf2 turnover pathway could not restore Nrf2 levels post RV-SA11 infection. Depletion of the Nrf2/HO-1 axis was subsequently found to be sensitive to proteasome inhibition with concurrent observation of increased K48-linked ubiquitination associated with Nrf2. Together, the present study describes robust downregulation of Nrf2-dependent cellular redox defense beyond initial hours of RV infection, justifying our previous observation of potent antirotaviral implications of Nrf2 agonists.
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Activation of PI3K, Akt, and ERK during early rotavirus infection leads to V-ATPase-dependent endosomal acidification required for uncoating. PLoS Pathog 2018; 14:e1006820. [PMID: 29352319 PMCID: PMC5792019 DOI: 10.1371/journal.ppat.1006820] [Citation(s) in RCA: 30] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Academic Contribution Register] [Received: 09/29/2017] [Revised: 01/31/2018] [Accepted: 12/15/2017] [Indexed: 11/19/2022] Open
Abstract
The cellular PI3K/Akt and/or MEK/ERK signaling pathways mediate the entry process or endosomal acidification during infection of many viruses. However, their roles in the early infection events of group A rotaviruses (RVAs) have remained elusive. Here, we show that late-penetration (L-P) human DS-1 and bovine NCDV RVA strains stimulate these signaling pathways very early in the infection. Inhibition of both signaling pathways significantly reduced production of viral progeny due to blockage of virus particles in the late endosome, indicating that neither of the two signaling pathways is involved in virus trafficking. However, immunoprecipitation assays using antibodies specific for pPI3K, pAkt, pERK and the subunit E of the V-ATPase co-immunoprecipitated the V-ATPase in complex with pPI3K, pAkt, and pERK. Moreover, Duolink proximity ligation assay revealed direct association of the subunit E of the V-ATPase with the molecules pPI3K, pAkt, and pERK, indicating that both signaling pathways are involved in V-ATPase-dependent endosomal acidification. Acidic replenishment of the medium restored uncoating of the RVA strains in cells pretreated with inhibitors specific for both signaling pathways, confirming the above results. Isolated components of the outer capsid proteins, expressed as VP4-VP8* and VP4-VP5* domains, and VP7, activated the PI3K/Akt and MEK/ERK pathways. Furthermore, psoralen-UV-inactivated RVA and CsCl-purified RVA triple-layered particles triggered activation of the PI3K/Akt and MEK/ERK pathways, confirming the above results. Our data demonstrate that multistep binding of outer capsid proteins of L-P RVA strains with cell surface receptors phosphorylates PI3K, Akt, and ERK, which in turn directly interact with the subunit E of the V-ATPase to acidify the late endosome for uncoating of RVAs. This study provides a better understanding of the RVA-host interaction during viral uncoating, which is of importance for the development of strategies aiming at controlling or preventing RVA infections. Viral particles must transport their genome into the cytoplasm or the nucleus of host cells to initiate successful infection. Knowledge of how viruses may pirate host cell signaling cascades or molecules to promote their own replication can facilitate the development of antiviral drugs. Group A rotavirus (RVA) is a major etiological agent of acute gastroenteritis in young children and the young of various mammals. RVA enters cells by a complex multistep process. However, the cellular signaling cascades or molecules that facilitate these processes are incompletely understood. Here, we demonstrate that infection with late-penetration RVA strains results in phosphorylation of PI3K, Akt, and ERK signaling molecules at an early stage of infection, a process mediated by the multistep binding of RVAs outer capsid proteins. Specific inhibitors for PI3K/Akt and MEK/ERK signaling pathways trap the viral particles in late endosome, and acidic replenishment restores and releases them. Moreover, the RVA-induced phosphorylated PI3K, Akt, and ERK directly interact with the subunit E of the V-ATPase proton pump, required for endosomal acidification and RVA uncoating. Understanding how RVA-induced early activation of cellular signaling molecules mediates the V-ATPase-dependent endosomal acidification required for uncoating of viral particles opens up opportunities for targeted interventions against rotavirus entry.
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Holloway G, Fleming FE, Coulson BS. MHC class I expression in intestinal cells is reduced by rotavirus infection and increased in bystander cells lacking rotavirus antigen. Sci Rep 2018; 8:67. [PMID: 29311575 PMCID: PMC5758578 DOI: 10.1038/s41598-017-18464-x] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Academic Contribution Register] [Received: 05/25/2017] [Accepted: 12/08/2017] [Indexed: 12/19/2022] Open
Abstract
Detection of viral infection by host cells leads to secretion of type I interferon, which induces antiviral gene expression. The class I major histocompatibility complex (MHCI) is required for viral antigen presentation and subsequent infected cell killing by cytotoxic T lymphocytes. STAT1 activation by interferon can induce NLRC5 expression, promoting MHCI expression. Rotavirus, an important pathogen, blocks interferon signalling through inhibition of STAT1 nuclear translocation. We assessed MHCI expression in HT-29 intestinal epithelial cells following rotavirus infection. MHCI levels were upregulated in a partially type I interferon-dependent manner in bystander cells lacking rotavirus antigen, but not in infected cells. MHCI and NLRC5 mRNA expression also was elevated in bystander, but not infected, cells, suggesting a transcriptional block in infected cells. STAT1 was activated in bystander and infected cells, but showed nuclear localisation in bystander cells only. Overall, the lack of MHCI upregulation in rotavirus-infected cells may be at least partially due to rotavirus blockade of interferon-induced STAT1 nuclear translocation. The reduced MHCI protein levels in infected cells support the existence of an additional, non-transcriptional mechanism that reduces MHCI expression. It is possible that rotavirus also may suppress MHCI expression in vivo, which might limit T cell-mediated killing of rotavirus-infected enterocytes.
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Affiliation(s)
- Gavan Holloway
- Department of Microbiology and Immunology, The University of Melbourne at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
| | - Fiona E Fleming
- Department of Microbiology and Immunology, The University of Melbourne at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia
| | - Barbara S Coulson
- Department of Microbiology and Immunology, The University of Melbourne at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria, Australia.
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Kindrachuk J. Selective inhibition of host cell signaling for rotavirus antivirals: PI3K/Akt/mTOR-mediated rotavirus pathogenesis. Virulence 2017; 9:5-8. [PMID: 28723236 PMCID: PMC5955445 DOI: 10.1080/21505594.2017.1356539] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Academic Contribution Register] [Indexed: 12/17/2022] Open
Affiliation(s)
- Jason Kindrachuk
- a Laboratory of Emerging and Re-Emerging Viruses , Department of Medical Microbiology, University of Manitoba , Winnipeg , MB , Canada
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Guerrero CA, Acosta O. Inflammatory and oxidative stress in rotavirus infection. World J Virol 2016; 5:38-62. [PMID: 27175349 PMCID: PMC4861870 DOI: 10.5501/wjv.v5.i2.38] [Citation(s) in RCA: 34] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Academic Contribution Register] [Received: 08/12/2015] [Revised: 10/23/2015] [Accepted: 01/29/2016] [Indexed: 02/05/2023] Open
Abstract
Rotaviruses are the single leading cause of life-threatening diarrhea affecting children under 5 years of age. Rotavirus entry into the host cell seems to occur by sequential interactions between virion proteins and various cell surface molecules. The entry mechanisms seem to involve the contribution of cellular molecules having binding, chaperoning and oxido-reducing activities. It appears to be that the receptor usage and tropism of rotaviruses is determined by the species, cell line and rotavirus strain. Rotaviruses have evolved functions which can antagonize the host innate immune response, whereas are able to induce endoplasmic reticulum (ER) stress, oxidative stress and inflammatory signaling. A networking between ER stress, inflammation and oxidative stress is suggested, in which release of calcium from the ER increases the generation of mitochondrial reactive oxygen species (ROS) leading to toxic accumulation of ROS within ER and mitochondria. Sustained ER stress potentially stimulates inflammatory response through unfolded protein response pathways. However, the detailed characterization of the molecular mechanisms underpinning these rotavirus-induced stressful conditions is still lacking. The signaling events triggered by host recognition of virus-associated molecular patterns offers an opportunity for the development of novel therapeutic strategies aimed at interfering with rotavirus infection. The use of N-acetylcysteine, non-steroidal anti-inflammatory drugs and PPARγ agonists to inhibit rotavirus infection opens a new way for treating the rotavirus-induced diarrhea and complementing vaccines.
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Moreno LY, Guerrero CA, Acosta O. Interacciones de las proteínas disulfuro isomerasa y de choque térmico Hsc70 con proteínas estructurales recombinantes purificadas de rotavirus. REVISTA COLOMBIANA DE BIOTECNOLOGÍA 2016. [DOI: 10.15446/rev.colomb.biote.v18n1.57714] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Academic Contribution Register] [Indexed: 11/09/2022] Open
Abstract
<p>Introducción. La entrada de rotavirus a las células parece estar mediado por interacciones secuenciales entre las proteínas estructurales virales y algunas moléculas de la superficie celular. Sin embargo, los mecanismos por los cuales el rotavirus infecta la célula diana aún no se comprenden bien. Existe alguna evidencia que muestra que las proteínas estructurales de rotavirus VP5* y VP8* interactúan con algunas moléculas de la superficie celular. La disponibilidad de las proteínas estructurales de rotavirus recombinantes en cantidad suficiente se ha convertido en un aspecto importante para la identificación de las interacciones específicas de los receptores virus-célula durante los eventos tempranos del proceso infeccioso. Objetivo. El propósito del presente trabajo es realizar un análisis de las interacciones entre las proteínas estructurales de rotavirus recombinante VP5*, VP8* y VP6, y las proteínas celulares Hsc70 y PDI utilizando sus versiones recombinantes purificadas. Materiales y métodos. Las proteínas recombinantes de rotavirus VP5* y VP8* y las proteínas recombinantes celulares Hsc70 y PDI se expresaron en E. BL21 (DE3), mientras que VP6 se expresó en células MA104 con virus vaccinia recombinante transfectada. La interacción entre el rotavirus y las proteínas celulares se estudió mediante ELISA, co-inmunoprecipitación y SDS-PAGE/ Western. Resultados. Las condiciones óptimas para la expresión de proteínas recombinantes se determinaron y se generaron anticuerpos contra ellas. Los resultados sugirieron que las proteínas virales rVP5* y rVP6 interactúan con Hsc70 y PDI in vitro. También se encontró que éstas proteínas virales recombinantes interactúan con Hsc70 en las balsas lipídicas (“Rafts”) en un cultivo celular. El tratamiento de las células, ya sea con DLP o rVP6 produjo significativamente la inhibición de la infección por rotavirus. Conclusión. Los resultados permiten concluir que rVP5 * y rVP6 interactúan con Hsc70 y PDI durante el proceso de la infección por rotavirus.</p>
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Kumar P, van den Hurk J, Ayalew LE, Gaba A, Tikoo SK. Proteomic analysis of purified turkey adenovirus 3 virions. Vet Res 2015; 46:79. [PMID: 26159706 PMCID: PMC4497381 DOI: 10.1186/s13567-015-0214-z] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Academic Contribution Register] [Received: 01/07/2015] [Accepted: 06/10/2015] [Indexed: 11/30/2022] Open
Abstract
Turkey adenovirus 3 (TAdV-3) causes high mortality and significant economic losses to the turkey industry. However, little is known about the molecular determinants required for viral replication and pathogenesis. Moreover, TAdV-3 does not grow well in cell culture, thus detailed structural studies of the infectious particle is particularly challenging. To develop a better understanding of virus-host interactions, we performed a comprehensive proteomic analysis of proteinase K treated purified TAdV-3 virions isolated from spleens of infected turkeys, by utilizing one-dimensional liquid chromatography mass spectrometry. Our analysis resulted in the identification of 13 viral proteins associated with TAdV-3 virions including a novel uncharacterized TaV3gp04 protein. Further, we detected 18 host proteins in purified virions, many of which are involved in cell-to cell spread, cytoskeleton dynamics and virus replication. Notably, seven of these host proteins have not yet been reported to be present in any other purified virus. In addition, five of these proteins are known antiviral host restriction factors. The availability of reagents allowed us to identify two cellular proteins (collagen alpha-1 (VI) chain and haemoglobin) in the purified TAdV-3 preparations. These results represent the first comprehensive proteomic profile of TAdV-3 and may provide information for illustrating TAdV-3 replication and pathogenesis.
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Affiliation(s)
- Pankaj Kumar
- Vaccine and Infectious Disease Organization -International Vaccine Center (VIDO- InterVac1), University of Saskatchewan, Saskatoon, S7N 5E3, SK, Canada.
| | - Jan van den Hurk
- Vaccine and Infectious Disease Organization -International Vaccine Center (VIDO- InterVac1), University of Saskatchewan, Saskatoon, S7N 5E3, SK, Canada.
| | - Lisanework E Ayalew
- Vaccine and Infectious Disease Organization -International Vaccine Center (VIDO- InterVac1), University of Saskatchewan, Saskatoon, S7N 5E3, SK, Canada.
| | - Amit Gaba
- Vaccine and Infectious Disease Organization -International Vaccine Center (VIDO- InterVac1), University of Saskatchewan, Saskatoon, S7N 5E3, SK, Canada.
| | - Suresh K Tikoo
- Vaccine and Infectious Disease Organization -International Vaccine Center (VIDO- InterVac1), University of Saskatchewan, Saskatoon, S7N 5E3, SK, Canada. .,Vaccinology & Immunotherapeutics program, School of Public Health, University of Saskatchewan, Saskatoon, S7N 5E5, SK, Canada.
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13
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Innate immune responses to rotavirus infection in macrophages depend on MAVS but involve neither the NLRP3 inflammasome nor JNK and p38 signaling pathways. Virus Res 2015; 208:89-97. [PMID: 26079065 DOI: 10.1016/j.virusres.2015.06.004] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Academic Contribution Register] [Received: 04/29/2015] [Revised: 06/03/2015] [Accepted: 06/04/2015] [Indexed: 12/24/2022]
Abstract
Rotavirus infection is a major cause of life-threatening infantile gastroenteritis. The innate immune system provides an immediate mechanism of suppressing viral replication and is necessary for an effective adaptive immune response. Innate immunity involves host recognition of viral infection and establishment of a powerful antiviral state through the expression of pro-inflammatory cytokines such as type-1 interferon (IFN). Macrophages, the front-line cells of innate immunity, produce IFN and other cytokines in response to viral infection. However, the role of macrophages during rotavirus infection is not well defined. We demonstrate here that RRV rotavirus triggers the production of proinflammatory cytokines from mouse bone marrow-derived macrophages. IFN and antiviral cytokine production was abolished in rotavirus-infected MAVS (-/-) macrophages. This indicates that rotavirus triggers innate immunity in macrophages through RIG-I and/or MDA5 viral recognition, and MAVS signaling is essential for cytokine responses in macrophages. Rotavirus induced IFN expression in both wild type and MDA5 (-/-) macrophages, showing that MDA5 is not essential for IFN secretion following infection, and RIG-I and MDA5 may act redundantly in promoting rotavirus recognition. Interestingly, rotavirus neither stimulated mitogen-activated protein kinases p38 and JNK nor activated the NLRP3 inflammasome, demonstrating that these components might not be involved in innate responses to rotavirus infection in macrophages. Our results indicate that rotavirus elicits intracellular signaling in macrophages, resulting in the induction of IFN and antiviral cytokines, and advance our understanding of the involvement of these cells in innate responses against rotavirus.
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Di Fiore IJM, Pane JA, Holloway G, Coulson BS. NSP1 of human rotaviruses commonly inhibits NF-κB signalling by inducing β-TrCP degradation. J Gen Virol 2015; 96:1768-76. [PMID: 25701827 DOI: 10.1099/vir.0.000093] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Academic Contribution Register] [Indexed: 12/21/2022] Open
Abstract
Rotavirus is a leading cause of severe gastroenteritis in infants worldwide. Rotavirus nonstructural protein 1 (NSP1) is a virulence factor that inhibits innate host immune responses. NSP1 from some rotaviruses targets host interferon response factors (IRFs), leading to inhibition of type I interferon expression. A few rotaviruses encode an NSP1 that inhibits the NF-κB pathway by targeting β-TrCP, a protein required for IκB degradation and NF-κB activation. Available evidence suggests that these NSP1 properties involve proteosomal degradation of target proteins. We show here that NSP1 from several human rotaviruses and porcine rotavirus CRW-8 inhibits the NF-κB pathway, but cannot degrade IRF3. Furthermore, β-TrCP levels were much reduced in cells infected with these rotaviruses. This provides strong evidence that β-TrCP degradation is required for NF-κB pathway inhibition by NSP1 and demonstrates the relevance of β-TrCP degradation to rotavirus infection. C-terminal regions of NSP1, including a serine-containing motif resembling the β-TrCP recognition motif of IκB, were required for NF-κB inhibition. CRW-8 infection of HT-29 intestinal epithelial cells induced significant levels of IFN-β and CCL5 but not IL-8. This contrasts with monkey rotavirus SA11-4F, whose NSP1 inhibits IRF3 but not NF-κB. Substantial amounts of IL-8 but not IFN-β or CCL5 were secreted from HT-29 cells infected with SA11-4F. Our results show that human rotaviruses commonly inhibit the NF-κB pathway by degrading β-TrCP and thus stabilizing IκB. They suggest that NSP1 plays an important role during human rotavirus infection by inhibiting the expression of NF-κB-dependent cytokines, such as IL-8.
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Affiliation(s)
- Izabel J M Di Fiore
- Department of Microbiology and Immunology, University of Melbourne at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria 3000, Australia
| | - Jessica A Pane
- Department of Microbiology and Immunology, University of Melbourne at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria 3000, Australia
| | - Gavan Holloway
- Department of Microbiology and Immunology, University of Melbourne at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria 3000, Australia
| | - Barbara S Coulson
- Department of Microbiology and Immunology, University of Melbourne at the Peter Doherty Institute for Infection and Immunity, Melbourne, Victoria 3000, Australia
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15
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Guerrero CA, Paula Pardo VR, Rafael Guerrero OA. Inhibition of rotavirus ECwt infection in ICR suckling mice by N-acetylcysteine, peroxisome proliferator-activated receptor gamma agonists and cyclooxygenase-2 inhibitors. Mem Inst Oswaldo Cruz 2014; 108:741-54. [PMID: 24037197 PMCID: PMC3970679 DOI: 10.1590/0074-0276108062013011] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Academic Contribution Register] [Received: 02/10/2013] [Accepted: 06/21/2013] [Indexed: 01/01/2023] Open
Abstract
Live attenuated vaccines have recently been introduced for preventing rotavirus
disease in children. However, alternative strategies for prevention and
treatment of rotavirus infection are needed mainly in developing countries where
low vaccine coverage occurs. In the present work, N-acetylcysteine (NAC),
ascorbic acid (AA), some nonsteroidal anti-inflammatory drugs (NSAIDs) and
peroxisome proliferator-activated receptor gamma (PPARγ) agonists were tested
for their ability to interfere with rotavirus ECwt infectivity as detected by
the percentage of viral antigen-positive cells of small intestinal villi
isolated from ECwt-infected ICR mice. Administration of 6 mg NAC/kg every 8 h
for three days following the first diarrhoeal episode reduced viral infectivity
by about 90%. Administration of AA, ibuprofen, diclofenac, pioglitazone or
rosiglitazone decreased viral infectivity by about 55%, 90%, 35%, 32% and 25%,
respectively. ECwt infection of mice increased expression of cyclooxygenase-2,
ERp57, Hsc70, NF-κB, Hsp70, protein disulphide isomerase (PDI) and PPARγ in
intestinal villus cells. NAC treatment of ECwt-infected mice reduced Hsc70 and
PDI expression to levels similar to those observed in villi from uninfected
control mice. The present results suggest that the drugs tested in the present
work could be assayed in preventing or treating rotaviral diarrhoea in children
and young animals.
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16
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Pane JA, Webster NL, Coulson BS. Rotavirus activates lymphocytes from non-obese diabetic mice by triggering toll-like receptor 7 signaling and interferon production in plasmacytoid dendritic cells. PLoS Pathog 2014; 10:e1003998. [PMID: 24676425 PMCID: PMC3968122 DOI: 10.1371/journal.ppat.1003998] [Citation(s) in RCA: 42] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Download PDF] [Figures] [Journal Information] [Subscribe] [Academic Contribution Register] [Received: 08/26/2013] [Accepted: 01/30/2014] [Indexed: 12/15/2022] Open
Abstract
It has been proposed that rotavirus infection promotes the progression of genetically-predisposed children to type 1 diabetes, a chronic autoimmune disease marked by infiltration of activated lymphocytes into pancreatic islets. Non-obese diabetic (NOD) mice provide a model for the human disease. Infection of adult NOD mice with rhesus monkey rotavirus (RRV) accelerates diabetes onset, without evidence of pancreatic infection. Rather, RRV spreads to the pancreatic and mesenteric lymph nodes where its association with antigen-presenting cells, including dendritic cells, induces cellular maturation. RRV infection increases levels of the class I major histocompatibility complex on B cells and proinflammatory cytokine expression by T cells at these sites. In autoimmunity-resistant mice and human mononuclear cells from blood, rotavirus-exposed plasmacytoid dendritic cells contribute to bystander polyclonal B cell activation through type I interferon expression. Here we tested the hypothesis that rotavirus induces bystander activation of lymphocytes from NOD mice by provoking dendritic cell activation and proinflammatory cytokine secretion. NOD mouse splenocytes were stimulated with rotavirus and assessed for activation by flow cytometry. This stimulation activated antigen-presenting cells and B cells independently of virus strain and replicative ability. Instead, activation depended on virus dose and was prevented by blockade of virus decapsidation, inhibition of endosomal acidification and interference with signaling through Toll-like receptor 7 and the type I interferon receptor. Plasmacytoid dendritic cells were more efficiently activated than conventional dendritic cells by RRV, and contributed to the activation of B and T cells, including islet-autoreactive CD8+ T cells. Thus, a double-stranded RNA virus can induce Toll-like receptor 7 signaling, resulting in lymphocyte activation. Our findings suggest that bystander activation mediated by type I interferon contributes to the lymphocyte activation observed following RRV infection of NOD mice, and may play a role in diabetes acceleration by rotavirus.
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Affiliation(s)
- Jessica A. Pane
- Department of Microbiology and Immunology, The University of Melbourne, Parkville, Victoria, Australia
| | - Nicole L. Webster
- Department of Microbiology and Immunology, The University of Melbourne, Parkville, Victoria, Australia
| | - Barbara S. Coulson
- Department of Microbiology and Immunology, The University of Melbourne, Parkville, Victoria, Australia
- * E-mail:
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Wang J, Hu G, Gao W, Xu L, Ning P, Zhang Y. Immortalized porcine intestinal epithelial cell cultures susceptible to porcine rotavirus infection. J Virol Methods 2014; 202:87-94. [PMID: 24642240 DOI: 10.1016/j.jviromet.2014.03.007] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Academic Contribution Register] [Received: 11/01/2013] [Revised: 02/13/2014] [Accepted: 03/04/2014] [Indexed: 11/17/2022]
Abstract
In vitro studies related to various viral pathogenesis in swine have been hampered by the lack of relevant porcine cell lines. The susceptibility to porcine rotavirus infection was evaluated by using a newly established porcine intestinal epithelial cell line. Immunohistochemical staining for cytokeratin confirmed that the cultured cells were epithelial cells. Measurement of cell viability and detection of infected cells confirmed that these epithelial cells were susceptible to porcine rotavirus infection. This study describes the cytopathic changes in cultured porcine intestinal epithelial cells during virus invasion. Following infection with porcine rotavirus, the cell cultures contained viral protein at 16 h post-infection as detected by direct immunofluorescence. The epithelial cell cultures provided competent target cells for studying host cell responses to porcine rotavirus and a homologous system for investigating the response of intestinal epithelial cells during viral infection.
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Affiliation(s)
- Jing Wang
- College of Veterinary Medicine, Northwest A&F University, Yangling 712100, Shaanxi, China
| | - Guangdong Hu
- College of Veterinary Medicine, Northwest A&F University, Yangling 712100, Shaanxi, China; Key Laboratory of Animal Biotechnology, Ministry of Agriculture, Northwest A&F University, Yangling 712100, Shaanxi, China
| | - Wanjun Gao
- College of Veterinary Medicine, Northwest A&F University, Yangling 712100, Shaanxi, China
| | - Lei Xu
- College of Veterinary Medicine, Northwest A&F University, Yangling 712100, Shaanxi, China
| | - Pengbo Ning
- College of Veterinary Medicine, Northwest A&F University, Yangling 712100, Shaanxi, China
| | - Yanming Zhang
- College of Veterinary Medicine, Northwest A&F University, Yangling 712100, Shaanxi, China.
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Relative roles of GM1 ganglioside, N-acylneuraminic acids, and α2β1 integrin in mediating rotavirus infection. J Virol 2014; 88:4558-71. [PMID: 24501414 DOI: 10.1128/jvi.03431-13] [Citation(s) in RCA: 42] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Academic Contribution Register] [Indexed: 12/22/2022] Open
Abstract
UNLABELLED N-acetyl- and N-glycolylneuraminic acids (Sia) and α2β1 integrin are frequently used by rotaviruses as cellular receptors through recognition by virion spike protein VP4. The VP4 subunit VP8*, derived from Wa rotavirus, binds the internal N-acetylneuraminic acid on ganglioside GM1. Wa infection is increased by enhanced internal Sia access following terminal Sia removal from main glycan chains with sialidase. The GM1 ligand cholera toxin B (CTB) reduces Wa infectivity. Here, we found sialidase treatment increased cellular GM1 availability and the infectivity of several other human (including RV-3) and animal rotaviruses, typically rendering them susceptible to methyl α-d-N-acetylneuraminide treatment, but did not alter α2β1 usage. CTB reduced the infectivity of these viruses. Aceramido-GM1 inhibited Wa and RV-3 infectivity in untreated and sialidase-treated cells, and GM1 supplementation increased their infectivity, demonstrating the importance of GM1 for infection. Wa recognition of α2β1 and internal Sia were at least partially independent. Rotavirus usage of GM1 was mapped to VP4 using virus reassortants, and RV-3 VP8* bound aceramido-GM1 by saturation transfer difference nuclear magnetic resonance (STD NMR). Most rotaviruses recognizing terminal Sia did not use GM1, including RRV. RRV VP8* interacted minimally with aceramido-GM1 by STD NMR. Unusually, TFR-41 rotavirus infectivity depended upon terminal Sia and GM1. Competition of CTB, Sia, and/or aceramido-GM1 with cell binding by VP8* from representative rotaviruses showed that rotavirus Sia and GM1 preferences resulted from VP8*-cell binding. Our major finding is that infection by human rotaviruses of commonly occurring VP4 serotypes involves VP8* binding to cell surface GM1 glycan, typically including the internal N-acetylneuraminic acid. IMPORTANCE Rotaviruses, the major cause of severe infantile gastroenteritis, recognize cell surface receptors through virus spike protein VP4. Several animal rotaviruses are known to bind sialic acids at the termini of main carbohydrate chains. Conversely, only a single human rotavirus is known to bind sialic acid. Interestingly, VP4 of this rotavirus bound to sialic acid that forms a branch on the main carbohydrate chain of the GM1 ganglioside. Here, we use several techniques to demonstrate that other human rotaviruses exhibit similar GM1 usage properties. Furthermore, binding by VP4 to cell surface GM1, involving branched sialic acid recognition, is shown to facilitate infection. In contrast, most animal rotaviruses that bind terminal sialic acids did not utilize GM1 for VP4 cell binding or infection. These studies support a significant role for GM1 in mediating host cell invasion by human rotaviruses.
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Diehl N, Schaal H. Make yourself at home: viral hijacking of the PI3K/Akt signaling pathway. Viruses 2013; 5:3192-212. [PMID: 24351799 PMCID: PMC3967167 DOI: 10.3390/v5123192] [Citation(s) in RCA: 149] [Impact Index Per Article: 12.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Download PDF] [Figures] [Journal Information] [Subscribe] [Academic Contribution Register] [Received: 10/29/2013] [Revised: 12/03/2013] [Accepted: 12/05/2013] [Indexed: 01/04/2023] Open
Abstract
As viruses do not possess genes encoding for proteins required for translation, energy metabolism or membrane biosynthesis, they are classified as obligatory intracellular parasites that depend on a host cell to replicate. This genome limitation forces them to gain control over cellular processes to ensure their successful propagation. A diverse spectrum of virally encoded proteins tackling a broad spectrum of cellular pathways during most steps of the viral life cycle ranging from the host cell entry to viral protein translation has evolved. Since the host cell PI3K/Akt signaling pathway plays a critical regulatory role in many cellular processes including RNA processing, translation, autophagy and apoptosis, many viruses, in widely varying ways, target it. This review focuses on a number of remarkable examples of viral strategies, which exploit the PI3K/Akt signaling pathway for effective viral replication.
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Affiliation(s)
| | - Heiner Schaal
- Universitätsklinikum Düsseldorf, Institut für Virologie, Universitätsstraße 1, Düsseldorf 40225, Germany.
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20
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Pane JA, Webster NL, Graham KL, Holloway G, Zufferey C, Coulson BS. Rotavirus acceleration of murine type 1 diabetes is associated with a T helper 1-dependent specific serum antibody response and virus effects in regional lymph nodes. Diabetologia 2013; 56:573-82. [PMID: 23238791 DOI: 10.1007/s00125-012-2798-4] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Academic Contribution Register] [Received: 08/20/2012] [Accepted: 11/16/2012] [Indexed: 01/16/2023]
Abstract
AIMS/HYPOTHESIS Rotavirus infection in at-risk children correlates with production of serum autoantibodies indicative of type 1 diabetes progression. Oral infection with rhesus monkey rotavirus (RRV) accelerates diabetes onset in mice. This relates to their rotavirus-specific serum antibody titre and local pro-inflammatory cytokine induction without pancreatic infection. Our aim was to further investigate the roles of serum antibodies and viral extra-intestinal spread in diabetes acceleration by rotavirus. METHODS Rotavirus-specific serum antibody production was detected by ELISA in diabetes-prone mice given either inactivated or low-dose RRV, in relation to their diabetes development. Serum anti-rotavirus antibody titres and infectious virus in lymph nodes were measured in mice given RRV or porcine rotavirus CRW-8. In lymph node cells, rotavirus antigen presence and immune activation were determined by flow cytometry, in conjunction with cytokine mRNA levels. RESULTS Acceleration of diabetes by RRV required virus replication, which correlated with antibody presence. CRW-8 induced similar specific total immunoglobulin and IgA titres to those induced by RRV, but did not accelerate diabetes. RRV alone elicited specific serum IgG antibodies with a T helper (Th)1 bias, spread to regional lymph nodes and activated antigen-presenting cells at these sites. RRV increased Th1-specific cytokine expression in pancreatic lymph nodes. Diabetes onset was more rapid in the RRV-infected mice with the greater Th1 bias. CONCLUSIONS/INTERPRETATION Acceleration of murine diabetes by rotavirus is virus strain-specific and associated with virus spread to regional lymph nodes, activation of antigen-presenting cells at these sites and induction of a Th1-dominated antibody and cytokine response.
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Affiliation(s)
- J A Pane
- Department of Microbiology and Immunology, Gate 11, Royal Parade, The University of Melbourne, Melbourne, Victoria 3010, Australia
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21
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He H, Mou Z, Li W, Fei L, Tang Y, Zhang J, Yan P, Chen Z, Yang X, Shen Z, Li J, Wu Y. Proteomic methods reveal cyclophilin a function as a host restriction factor against rotavirus infection. Proteomics 2013; 13:1121-32. [DOI: 10.1002/pmic.201100579] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Academic Contribution Register] [Received: 11/04/2011] [Revised: 08/28/2012] [Accepted: 12/17/2012] [Indexed: 12/22/2022]
Affiliation(s)
- Haiyang He
- Institute of Immunology; Third Military Medical University; Chongqing P. R. China
| | - Zhirong Mou
- Institute of Immunology; Third Military Medical University; Chongqing P. R. China
| | - Wanling Li
- Institute of Immunology; Third Military Medical University; Chongqing P. R. China
| | - Lei Fei
- Institute of Immunology; Third Military Medical University; Chongqing P. R. China
| | - Yan Tang
- Institute of Immunology; Third Military Medical University; Chongqing P. R. China
| | - Ji Zhang
- Institute of Immunology; Third Military Medical University; Chongqing P. R. China
| | - Ping Yan
- Southwest Hospital; Third Military Medical University; Chongqing P. R. China
| | - Zhengqiong Chen
- Xinqiao Hospital; Third Military Medical University; Chongqing P. R. China
| | - Xia Yang
- Institute of Immunology; Third Military Medical University; Chongqing P. R. China
| | - Zigang Shen
- Institute of Immunology; Third Military Medical University; Chongqing P. R. China
| | - Jintao Li
- Institute of Immunology; Third Military Medical University; Chongqing P. R. China
| | - Yuzhang Wu
- Institute of Immunology; Third Military Medical University; Chongqing P. R. China
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Ye Y, Yan G, Luo Y, Tong T, Liu X, Xin C, Liao M, Fan H. Quantitative Proteomics by Amino Acid Labeling in Foot-and-Mouth Disease Virus (FMDV)-Infected Cells. J Proteome Res 2012; 12:363-77. [DOI: 10.1021/pr300611e] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Academic Contribution Register] [Indexed: 01/06/2023]
Affiliation(s)
- Yu Ye
- Key Laboratory of Animal Vaccine
Development, Ministry of Agriculture, Guangzhou
510642, China
- College of
Veterinary Medicine, South China Agricultural University, Guangzhou 510642, China
- National and Regional Joint Engineering Laboratory for Medicament of Zoonosis Prevention and Control, Guangzhou
510642, China
| | - Guangrong Yan
- Institute of Life and Health Engineering and National Engineering
and Research Center for Genetic Medicine, Jinan University, Guangzhou 510632, China
| | - Yongwen Luo
- College of
Veterinary Medicine, South China Agricultural University, Guangzhou 510642, China
| | - Tiezhu Tong
- Huizhou Entry-Exit Inspection and Quarantine Bureau, Huizhou 516001, China
| | - Xiangtao Liu
- State
Key Laboratory of Veterinary Etiologic Biology, Lanzhou Veterinary
Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou 730046, China
| | - Chaoan Xin
- Key Laboratory of Animal Vaccine
Development, Ministry of Agriculture, Guangzhou
510642, China
- College of
Veterinary Medicine, South China Agricultural University, Guangzhou 510642, China
- National and Regional Joint Engineering Laboratory for Medicament of Zoonosis Prevention and Control, Guangzhou
510642, China
| | - Ming Liao
- Key Laboratory of Animal Vaccine
Development, Ministry of Agriculture, Guangzhou
510642, China
- College of
Veterinary Medicine, South China Agricultural University, Guangzhou 510642, China
- National and Regional Joint Engineering Laboratory for Medicament of Zoonosis Prevention and Control, Guangzhou
510642, China
| | - Huiying Fan
- Key Laboratory of Animal Vaccine
Development, Ministry of Agriculture, Guangzhou
510642, China
- College of
Veterinary Medicine, South China Agricultural University, Guangzhou 510642, China
- National and Regional Joint Engineering Laboratory for Medicament of Zoonosis Prevention and Control, Guangzhou
510642, China
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23
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He H, Zhou D, Fan W, Fu X, Zhang J, Shen Z, Li J, Li J, Wu Y. Cyclophilin A inhibits rotavirus replication by facilitating host IFN-I production. Biochem Biophys Res Commun 2012; 422:664-9. [DOI: 10.1016/j.bbrc.2012.05.050] [Citation(s) in RCA: 25] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Academic Contribution Register] [Received: 05/01/2012] [Accepted: 05/09/2012] [Indexed: 10/28/2022]
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Cebo C, Martin P. Inter-species comparison of milk fat globule membrane proteins highlights the molecular diversity of lactadherin. Int Dairy J 2012. [DOI: 10.1016/j.idairyj.2011.09.017] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Academic Contribution Register] [Indexed: 12/24/2022]
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25
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Hardy ME, Hendricks JM, Paulson JM, Faunce NR. 18β-glycyrrhetinic acid inhibits rotavirus replication in culture. Virol J 2012; 9:96. [PMID: 22616823 PMCID: PMC3478227 DOI: 10.1186/1743-422x-9-96] [Citation(s) in RCA: 62] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Academic Contribution Register] [Received: 12/26/2011] [Accepted: 05/04/2012] [Indexed: 02/22/2023] Open
Abstract
BACKGROUND Glycyrrhizin (GA) and primary metabolite 18β-glycyrrhetinic acid (GRA) are pharmacologically active components of the medicinal licorice root, and both have been shown to have antiviral and immunomodulatory properties. Although these properties are well established, the mechanisms of action are not completely understood. In this study, GA and GRA were tested for the ability to inhibit rotavirus replication in cell culture, toward a long term goal of discovering natural compounds that may complement existing vaccines. METHODS Epithelial cells were treated with GA or GRA various times pre- or post-infection and virus yields were measured by immunofluorescent focus assay. Levels of viral proteins VP2, VP6, and NSP2 in GRA treated cells were measured by immunoblot to determine if there was an effect of GRA treatment on the accumulation of viral protein. RESULTS GRA treatment reduced rotavirus yields by 99% when added to infected cultures post-- virus adsorption, whereas virus yields in GA treated cultures were similar to mock treated controls. Time of addition experiments indicated that GRA-mediated replication inhibition likely occurs at a step or steps subsequent to virus entry. The amounts of VP2, VP6 and NSP2 were substantially reduced when GRA was added to cultures up to two hours post-entry. CONCLUSIONS GRA, but not GA, has significant antiviral activity against rotavirus replication in vitro, and studies to determine whether GRA attenuates rotavirus replication in vivo are underway.
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Affiliation(s)
- Michele E Hardy
- Immunology and Infectious Diseases, Montana State University, PO Box 173610, Bozeman, MT 59718, USA
| | - Jay M Hendricks
- Immunology and Infectious Diseases, Montana State University, PO Box 173610, Bozeman, MT 59718, USA
| | - Jeana M Paulson
- Immunology and Infectious Diseases, Montana State University, PO Box 173610, Bozeman, MT 59718, USA
| | - Nicholas R Faunce
- Immunology and Infectious Diseases, Montana State University, PO Box 173610, Bozeman, MT 59718, USA
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Steukers L, Glorieux S, Vandekerckhove AP, Favoreel HW, Nauwynck HJ. Diverse microbial interactions with the basement membrane barrier. Trends Microbiol 2012; 20:147-55. [PMID: 22300759 PMCID: PMC7127156 DOI: 10.1016/j.tim.2012.01.001] [Citation(s) in RCA: 27] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Academic Contribution Register] [Received: 08/31/2011] [Revised: 12/20/2011] [Accepted: 01/02/2012] [Indexed: 11/01/2022]
Abstract
During primary contact with susceptible hosts, microorganisms face an array of barriers that thwart their invasion process. Passage through the basement membrane (BM), a 50-100-nm-thick crucial barrier underlying epithelia and endothelia, is a prerequisite for successful host invasion. Such passage allows pathogens to reach nerve endings or blood vessels in the stroma and to facilitate spread to internal organs. During evolution, several pathogens have developed different mechanisms to cross this dense matrix of sheet-like proteins. To breach the BM, some microorganisms have developed independent mechanisms, others hijack host cells that are able to transverse the BM (e.g. leukocytes and dendritic cells) and oncogenic microorganisms might even trigger metastatic processes in epithelial cells to penetrate the underlying BM.
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Affiliation(s)
- Lennert Steukers
- Department of Virology, Parasitology and Immunology, Faculty of Veterinary Medicine, Ghent University, Salisburylaan 133, B-9820 Merelbeke, Belgium.
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Dunn EF, Connor JH. HijAkt: The PI3K/Akt pathway in virus replication and pathogenesis. PROGRESS IN MOLECULAR BIOLOGY AND TRANSLATIONAL SCIENCE 2012; 106:223-50. [PMID: 22340720 PMCID: PMC7149925 DOI: 10.1016/b978-0-12-396456-4.00002-x] [Citation(s) in RCA: 64] [Impact Index Per Article: 4.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Academic Contribution Register] [Indexed: 12/13/2022]
Abstract
As obligate parasites of cellular processes, viruses must take over cellular macromolecular machinery. It is also becoming clear that viruses routinely control intracellular signaling pathways through the direct or indirect control of kinases and phosphatases. This control of cellular phosphoproteins is important to promote a variety of viral processes, from control of entry to nuclear function to the stimulation of viral protein synthesis. This review focuses on the takeover of the cellular phosphatidylinositol-3-kinase (PI3K)/Akt signaling pathway by a variety of retroviruses, DNA viruses, and RNA viruses, highlighting the functions ascribed to virus activation of PI3K and Akt activity. This review also describes the role that the PI3K/Akt pathway plays in the host response, noting that it that can trigger anti- as well as proviral functions.
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Affiliation(s)
- Ewan F Dunn
- Department of Microbiology, Boston University School of Medicine, Boston, Massachusetts, USA
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Abstract
RNA viruses are the causative agents of severe diseases in vertebrates. Upon viral infection, various intracellular signaling pathways are induced within the infected cells. While most of the different cellular signaling responses are initiated as antiviral defense mechanisms to counteract invading pathogens, they may also be exploited by viruses to support their replication. Recently, PI3K has been added to the growing list of signaling factors and pathways that are activated upon viral infections and regulate the replication process. Here, the current knowledge on RNA virus-induced PI3K-regulated signaling processes and how the pathogens take advantage of these activities within the infected cells is summarized.
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Affiliation(s)
- Christina Ehrhardt
- Institute of Molecular Virology (IMV), ZMBE, Westfaelische-Wilhelms-University, Von Esmarch-Str. 56, D-48149 Münster, Germany
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Fleming FE, Graham KL, Takada Y, Coulson BS. Determinants of the specificity of rotavirus interactions with the alpha2beta1 integrin. J Biol Chem 2010; 286:6165-74. [PMID: 21138834 DOI: 10.1074/jbc.m110.142992] [Citation(s) in RCA: 25] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Academic Contribution Register] [Indexed: 12/15/2022] Open
Abstract
The human α2β1 integrin binds collagen and acts as a cellular receptor for rotaviruses and human echovirus 1. These ligands require the inserted (I) domain within the α2 subunit of α2β1 for binding. Previous studies have identified the binding sites for collagen and echovirus 1 in the α2 I domain. We used CHO cells expressing mutated α2β1 to identify amino acids involved in binding to human and animal rotaviruses. Residues where mutation affected rotavirus binding were located in several exposed loops and adjacent regions of the α2 I domain. Binding by all rotaviruses was eliminated by mutations in the activation-responsive αC-α6 and αF helices. This is a novel feature that distinguishes rotavirus from other α2β1 ligands. Mutation of residues that co-ordinate the metal ion (Ser-153, Thr-221, and Glu-256 in α2 and Asp-130 in β1) and nearby amino acids (Ser-154, Gln-215, and Asp-219) also inhibited rotavirus binding. The importance of most of these residues was greatest for binding by human rotaviruses. These mutations inhibit collagen binding to α2β1 (apart from Glu-256) but do not affect echovirus binding. Overall, residues where mutation affected both rotavirus and collagen recognition are located at one side of the metal ion-dependent adhesion site, whereas those important for collagen alone cluster nearby. Mutations eliminating rotavirus and echovirus binding are distinct, consistent with the respective preference of these viruses for activated or inactive α2β1. In contrast, rotavirus and collagen utilize activated α2β1 and show an overlap in α2β1 residues important for binding.
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Affiliation(s)
- Fiona E Fleming
- Department of Microbiology and Immunology, The University of Melbourne, Parkville, Victoria 3010, Australia
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30
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Guerrero CA, Santana AY, Acosta O. Mouse intestinal villi as a model system for studies of rotavirus infection. J Virol Methods 2010; 168:22-30. [DOI: 10.1016/j.jviromet.2010.04.010] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Academic Contribution Register] [Received: 09/04/2009] [Revised: 03/29/2010] [Accepted: 04/12/2010] [Indexed: 12/24/2022]
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Integrin alpha4beta7 is downregulated on the surfaces of simian immunodeficiency virus SIVmac239-infected cells. J Virol 2010; 84:6344-51. [PMID: 20410278 DOI: 10.1128/jvi.00430-10] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Academic Contribution Register] [Indexed: 11/20/2022] Open
Abstract
Simian immunodeficiency virus (SIV) and human immunodeficiency virus (HIV) infection results in an early and enduring depletion of intestinal CD4(+) T cells. SIV and HIV bind integrin alpha4beta7, thereby facilitating infection of lymphocytes that home to the gut-associated lymphoid tissue (GALT). Using an ex vivo flow cytometry assay, we found that SIVmac239-infected cells expressed significantly lower levels of integrin alpha4beta7 than did uninfected cells. This finding suggested a potential viral effect on integrin alpha4beta7 expression. Using an in vitro model, we confirmed that integrin alpha4beta7 was downregulated on the surfaces of SIVmac239-infected cells. Further, modulation of integrin alpha4beta7 was dependent on de novo synthesis of viral proteins, but neither cell death, the release of a soluble factor, nor a change in activation state was involved. Downregulation of integrin alpha4beta7 may have an unappreciated role in the CD4 depletion of the mucosal-associated lymphoid compartments, susceptibility to superinfection, and/or immune evasion.
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Halasz P, Holloway G, Coulson BS. Death mechanisms in epithelial cells following rotavirus infection, exposure to inactivated rotavirus or genome transfection. J Gen Virol 2010; 91:2007-2018. [PMID: 20392902 DOI: 10.1099/vir.0.018275-0] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Academic Contribution Register] [Indexed: 12/14/2022] Open
Abstract
Intestinal epithelial cell death following rotavirus infection is associated with villus atrophy and gastroenteritis. Roles for both apoptosis and necrosis in cytocidal activity within rotavirus-infected epithelial cells have been proposed. Additionally, inactivated rotavirus has been reported to induce diarrhoea in infant mice. We further examined the death mechanisms induced in epithelial cell lines following rotavirus infection or inactivated rotavirus exposure. Monolayer integrity changes in MA104, HT-29 and partially differentiated Caco-2 cells following inactivated rotavirus exposure or RRV or CRW-8 rotavirus infection paralleled cell metabolic activity and viability reductions. MA104 cell exposure to rotavirus dsRNA also altered monolayer integrity. Inactivated rotaviruses induced delayed cell function losses that were unrelated to apoptosis. Phosphatidylserine externalization, indicating early apoptosis, occurred in MA104 and HT-29 but not in partially differentiated Caco-2 cells by 11 h after infection. Rotavirus activation of phosphatidylinositol 3-kinase partially protected MA104 and HT-29 cells from early apoptosis. In contrast, activation of the stress-activated protein kinase JNK by rotavirus did not influence apoptosis induction in these cells. RRV infection produced DNA fragmentation, indicating late-stage apoptosis, in fully differentiated Caco-2 cells only. These studies show that the apoptosis initiation and cell death mechanism induced by rotavirus infection depend on cell type and degree of differentiation. Early stage apoptosis resulting from rotavirus infection is probably counter-balanced by virus-induced phosphatidylinositol 3-kinase activation. The ability of inactivated rotaviruses and rotavirus dsRNA to perturb monolayer integrity supports a potential role for these rotavirus components in disease pathogenesis.
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Affiliation(s)
- Peter Halasz
- Department of Microbiology and Immunology, The University of Melbourne, VIC 3010, Australia
| | - Gavan Holloway
- Department of Microbiology and Immunology, The University of Melbourne, VIC 3010, Australia
| | - Barbara S Coulson
- Department of Microbiology and Immunology, The University of Melbourne, VIC 3010, Australia
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Rotavirus nonstructural protein 1 suppresses virus-induced cellular apoptosis to facilitate viral growth by activating the cell survival pathways during early stages of infection. J Virol 2010; 84:6834-45. [PMID: 20392855 DOI: 10.1128/jvi.00225-10] [Citation(s) in RCA: 77] [Impact Index Per Article: 5.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Academic Contribution Register] [Indexed: 02/07/2023] Open
Abstract
Following virus infection, one of the cellular responses to limit the virus spread is induction of apoptosis. In the present study, we report role of rotavirus nonstructural protein 1 (NSP1) in regulating apoptosis by activating prosurvival pathways such as phosphatidylinositol 3-kinase (PI3K)/Akt and NF-kappaB (nuclear factor kappaB) during early hours of infections (2 to 8 hpi). The NSP1 mutant strain A5-16 induces weak and transient activation of Akt (protein kinase B) and p65 NF-kappaB compared to the isogenic wild-type strain A5-13 in MA104 or HT29 cells. The weak NF-kappaB promoter activity or Akt phosphorylation after A5-16 infection could be complemented in cells transfected with plasmid expressing NSP1 after infection with the rotavirus A5-16 strain. In cells either infected with A5-13 or transfected with pcD-NSP1, coimmunoprecipitation of NSP1 with phosphoinositide 3-kinase (PI3K) was observed, indicating that strong activation of PI3K/Akt could be due to its interaction with NSP1. In addition, after infection with same multiplicity of infection, A5-16 showed reduced number of viral particles compared to the A5-13 strain at the end of the replication cycle. A lower growth rate could be due to weak induction of PI3K/Akt and NF-kappaB, since the A5-13 strain also showed reduced growth in the presence of PI3K or NF-kappaB inhibitors. This effect was interferon independent; however, it was partly due to significantly higher caspase-3 activity, poly-ADP ribose polymerase (PARP) cleavage, and apoptosis during earlier stages of infection with the NSP1 mutant. Thus, our data suggest that NSP1 positively supports rotavirus growth by suppression of premature apoptosis for improved virus growth after infection.
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A dual effect of porcine reproductive and respiratory syndrome virus replication on the phosphatidylinositol-3-kinase-dependent Akt pathway. Arch Virol 2010; 155:571-5. [PMID: 20213282 DOI: 10.1007/s00705-010-0611-6] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Academic Contribution Register] [Received: 09/04/2009] [Accepted: 01/08/2010] [Indexed: 01/19/2023]
Abstract
We have recently shown that porcine reproductive and respiratory syndrome virus (PRRSV) can undergo a productive replication in porcine monocyte-derived dendritic cells (Mo-DCs). Here, we further demonstrate that PRRSV activates the host's phosphatidylinositol-3-kinase (PI3K)-dependent Akt pathway (PI3K/Akt) to facilitate its replication in Mo-DCs at 90 min and 4 h after infection. Inhibition of PI3K/Akt by treatment with a PI3K-specific inhibitor (LY294002) prior to PRRSV infection reduced virus replication. Furthermore, inhibition of PI3K/Akt by LY294002 at 90 min and 8 h after virus infection still significantly reduced virus production, suggesting that virus replication may be dependent on the activation of PI3K/Akt. Interestingly, PRRSV inhibited PI3K/Akt at 12 h after infection. Heat-inactivated virus failed to inhibit PI3K/Akt, indicating that virus replication is essential for this inhibition. Overall, PRRSV replication exhibits a dual effect on the PI3K/Akt pathway in which both time-dependent activation and inhibition of PI3K/Akt are observed.
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Dutta D, Bagchi P, Chatterjee A, Nayak MK, Mukherjee A, Chattopadhyay S, Nagashima S, Kobayashi N, Komoto S, Taniguchi K, Chawla-Sarkar M. The molecular chaperone heat shock protein-90 positively regulates rotavirus infectionx. Virology 2009; 391:325-33. [PMID: 19628238 DOI: 10.1016/j.virol.2009.06.044] [Citation(s) in RCA: 49] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Academic Contribution Register] [Received: 05/20/2009] [Revised: 06/07/2009] [Accepted: 06/26/2009] [Indexed: 11/17/2022]
Abstract
Rotaviruses are the major cause of severe dehydrating gastroenteritis in children worldwide. In this study, we report a positive role of cellular chaperone Hsp90 during rotavirus infection. A highly specific Hsp90 inhibitor, 17-allylamono-demethoxygeldanamycin (17-AAG) was used to delineate the functional role of Hsp90. In MA104 cells treated with 17-AAG after viral adsorption, replication of simian (SA11) or human (KU) strains was attenuated as assessed by quantitating both plaque forming units and expression of viral genes. Phosphorylation of Akt and NFkappaB observed 2-4 hpi with SA11, was strongly inhibited in the presence of 17-AAG. Direct Hsp90-Akt interaction in virus infected cells was also reduced in the presence of 17-AAG. Anti-rotaviral effects of 17-AAG were due to inhibition of activation of Akt that was confirmed since, PI3K/Akt inhibitors attenuated rotavirus growth significantly. Thus, Hsp90 regulates rotavirus by modulating cellular signaling proteins. The results highlight the importance of cellular proteins during rotavirus infection and the possibility of targeting cellular chaperones for developing new anti-rotaviral strategies.
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Affiliation(s)
- Dipanjan Dutta
- Division of Virology, National Institute of Cholera and Enteric Diseases, P-33, C.I.T. Road, Scheme-XM, Beliaghata, Kolkata-700010, West Bengal, India
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Wang H, Ma J, Ruan L, Xu X. Cloning of a centaurin-alpha1 like gene MjCent involved in WSSV infection from shrimp Marsupeneaus japonicus. FISH & SHELLFISH IMMUNOLOGY 2009; 26:279-284. [PMID: 19073266 DOI: 10.1016/j.fsi.2008.10.016] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Academic Contribution Register] [Received: 09/21/2008] [Revised: 10/22/2008] [Accepted: 10/28/2008] [Indexed: 05/27/2023]
Abstract
Centaurin-alpha1 specifically binds phosphatidylinositol 3,4,5-trisphosphate (PI (3,4,5)P3) and is a GTPase-activating protein (GAP) of ADP-ribosylation factor (ARF6). It actively engages in phosphatidylinositol 3-kinase (PI3-K) mediated cell signal transduction. Here, for the first time, we have identified a virus related centaurin-alpha1 homologue named MjCent from the shrimp, Marsupeneaus japonicus, an economically important crustacean in the aquaculture industry. MjCent has one conserved ArfGAP and two Pleckstrin homology domains (PH domains). As shown by RT-PCR and immunofluorescence, MjCent appeared in every tissue examined and was localized mainly in the cell cytoplasm. Further investigation with real-time quantitative PCR showed that MjCent was significantly up-regulated during white spot syndrome virus (WSSV) infection, but notably decreased in virus-resistant shrimps. This suggests a close relationship between MjCent and WSSV invasion and host defense of the shrimp, M. japonicus.
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Affiliation(s)
- Huifen Wang
- Key Laboratory of Marine Biogenetic Resources, Third Institute of Oceanography, State Oceanic Administration, Xiamen, 361005, PR China
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Saeed MF, Kolokoltsov AA, Freiberg AN, Holbrook MR, Davey RA. Phosphoinositide-3 kinase-Akt pathway controls cellular entry of Ebola virus. PLoS Pathog 2008; 4:e1000141. [PMID: 18769720 PMCID: PMC2516934 DOI: 10.1371/journal.ppat.1000141] [Citation(s) in RCA: 155] [Impact Index Per Article: 9.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Download PDF] [Figures] [Journal Information] [Subscribe] [Academic Contribution Register] [Received: 02/21/2008] [Accepted: 08/01/2008] [Indexed: 11/18/2022] Open
Abstract
The phosphoinositide-3 kinase (PI3K) pathway regulates diverse cellular activities related to cell growth, migration, survival, and vesicular trafficking. It is known that Ebola virus requires endocytosis to establish an infection. However, the cellular signals that mediate this uptake were unknown for Ebola virus as well as many other viruses. Here, the involvement of PI3K in Ebola virus entry was studied. A novel and critical role of the PI3K signaling pathway was demonstrated in cell entry of Zaire Ebola virus (ZEBOV). Inhibitors of PI3K and Akt significantly reduced infection by ZEBOV at an early step during the replication cycle. Furthermore, phosphorylation of Akt-1 was induced shortly after exposure of cells to radiation-inactivated ZEBOV, indicating that the virus actively induces the PI3K pathway and that replication was not required for this induction. Subsequent use of pseudotyped Ebola virus and/or Ebola virus-like particles, in a novel virus entry assay, provided evidence that activity of PI3K/Akt is required at the virus entry step. Class 1A PI3Ks appear to play a predominant role in regulating ZEBOV entry, and Rac1 is a key downstream effector in this regulatory cascade. Confocal imaging of fluorescently labeled ZEBOV indicated that inhibition of PI3K, Akt, or Rac1 disrupted normal uptake of virus particles into cells and resulted in aberrant accumulation of virus into a cytosolic compartment that was non-permissive for membrane fusion. We conclude that PI3K-mediated signaling plays an important role in regulating vesicular trafficking of ZEBOV necessary for cell entry. Disruption of this signaling leads to inappropriate trafficking within the cell and a block in steps leading to membrane fusion. These findings extend our current understanding of Ebola virus entry mechanism and may help in devising useful new strategies for treatment of Ebola virus infection.
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Affiliation(s)
- Mohammad F. Saeed
- Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, Texas, United States of America
- Western Regional Center of Excellence in Biodefense and Emerging Infectious Diseases Research, University of Texas Medical Branch, Galveston, Texas, United States of America
| | - Andrey A. Kolokoltsov
- Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, Texas, United States of America
| | - Alexander N. Freiberg
- Western Regional Center of Excellence in Biodefense and Emerging Infectious Diseases Research, University of Texas Medical Branch, Galveston, Texas, United States of America
- Department of Pathology, University of Texas Medical Branch, Galveston, Texas, United States of America
| | - Michael R. Holbrook
- Western Regional Center of Excellence in Biodefense and Emerging Infectious Diseases Research, University of Texas Medical Branch, Galveston, Texas, United States of America
- Department of Pathology, University of Texas Medical Branch, Galveston, Texas, United States of America
- Institute of Human Infection and Immunity, University of Texas Medical Branch, Galveston, Texas, United States of America
| | - Robert A. Davey
- Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, Texas, United States of America
- Western Regional Center of Excellence in Biodefense and Emerging Infectious Diseases Research, University of Texas Medical Branch, Galveston, Texas, United States of America
- Institute of Human Infection and Immunity, University of Texas Medical Branch, Galveston, Texas, United States of America
- * E-mail:
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Integrins alpha1beta1 and alpha2beta1 are receptors for the rotavirus enterotoxin. Proc Natl Acad Sci U S A 2008; 105:8811-8. [PMID: 18587047 DOI: 10.1073/pnas.0803934105] [Citation(s) in RCA: 63] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Academic Contribution Register] [Indexed: 02/06/2023] Open
Abstract
Rotavirus NSP4 is a viral enterotoxin capable of causing diarrhea in neonatal mice. This process is initiated by the binding of extracellular NSP4 to target molecule(s) on the cell surface that triggers a signaling cascade leading to diarrhea. We now report that the integrins alpha1beta1 and alpha2beta1 are receptors for NSP4. NSP4 specifically binds to the alpha1 and alpha2 I domains with apparent K(d) = 1-2.7 muM. Binding is mediated by the I domain metal ion-dependent adhesion site motif, requires Mg(2+) or Mn(2+), is abolished with EDTA, and an NSP4 point mutant, E(120)A, fails to bind alpha2 integrin I domain. NSP4 has two distinct integrin interaction domains. NSP4 amino acids 114-130 are essential for binding to the I domain, and NSP4 peptide 114-135 blocks binding of the natural ligand, collagen I, to integrin alpha2. NSP4 amino acids 131-140 are not associated with the initial binding to the I domain, but elicit signaling that leads to the spreading of attached C2C12-alpha2 cells, mouse myoblast cells stably expressing the human alpha2 integrin. NSP4 colocalizes with integrin alpha2 on the basolateral surface of rotavirus-infected polarized intestinal epithelial (Caco-2) cells as well as surrounding noninfected cells. NSP4 mutants that fail to bind or signal through integrin alpha2 were attenuated in diarrhea induction in neonatal mice. These results indicate that NSP4 interaction with integrin alpha1 and alpha2 is an important component of enterotoxin function and rotavirus pathogenesis, further distinguishing this viral virulence factor from other microbial enterotoxins.
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Abstract
Infection modulates type 1 diabetes, a common autoimmune disease characterized by the destruction of insulin-producing islet beta cells in the pancreas. Childhood rotavirus infections have been associated with exacerbations in islet autoimmunity. Nonobese diabetic (NOD) mice develop lymphocytic islet infiltration (insulitis) and then clinical diabetes, whereas NOD8.3 TCR mice, transgenic for a T-cell receptor (TCR) specific for an important islet autoantigen, show more rapid diabetes onset. Oral infection of infant NOD mice with the monkey rotavirus strain RRV delays diabetes development. Here, the effect of RRV infection on diabetes development once insulitis is established was determined. NOD and NOD8.3 TCR mice were inoculated with RRV aged > or = 12 and 5 weeks, respectively. Diabetes onset was significantly accelerated in both models (P < 0.024), although RRV infection was asymptomatic and confined to the intestine. The degree of diabetes acceleration was related to the serum antibody titer to RRV. RRV-infected NOD mice showed a possible trend toward increased insulitis development. Infected males showed increased CD8(+) T-cell proportions in islets. Levels of beta-cell major histocompatibility complex class I expression and islet tumor necrosis factor alpha mRNA were elevated in at least one model. NOD mouse exposure to mouse rotavirus in a natural experiment also accelerated diabetes. Thus, rotavirus infection after beta-cell autoimmunity is established affects insulitis and exacerbates diabetes. A possible mechanism involves increased exposure of beta cells to immune recognition and activation of autoreactive T cells by proinflammatory cytokines. The timing of infection relative to mouse age and degree of insulitis determines whether diabetes onset is delayed, unaltered, or accelerated.
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