Life as a Vector of Dengue Virus: The Antioxidant Strategy of Mosquito Cells to Survive Viral Infection

Dengue fever is a mosquito-borne viral disease of increasing global importance. The disease has caused heavy burdens due to frequent outbreaks in tropical and subtropical areas of the world. The dengue virus (DENV) is generally transmitted between human hosts via the bite of a mosquito vector, primarily Aedes aegypti and Ae. albopictus as a minor species. It is known that the virus needs to alternately infect mosquito and human cells. DENV-induced cell death is relevant to the pathogenesis in humans as infected cells undergo apoptosis. In contrast, mosquito cells mostly survive the infection; this allows infected mosquitoes to remain healthy enough to serve as an efficient vector in nature. Overexpression of antioxidant genes such as superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), glutathione S-transferase (GST), glutaredoxin (Grx), thioredoxin (Trx), and protein disulfide isomerase (PDI) have been detected in DENV2-infected mosquito cells. Additional antioxidants, including GST, eukaryotic translation initiation factor 5A (eIF5a), and p53 isoform 2 (p53-2), and perhaps some others, are also involved in creating an intracellular environment suitable for cell replication and viral infection. Antiapoptotic effects involving inhibitor of apoptosis (IAP) upregulation and subsequent elevation of caspase-9 and caspase-3 activities also play crucial roles in the ability of mosquito cells to survive DENV infection. This article focused on the effects of intracellular responses in mosquito cells to infection primarily by DENVs. It may provide more information to better understand virus/cell interactions that can possibly elucidate the evolutionary pathway that led to the mosquito becoming a vector.

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Upregulation of a novel eukaryotic translation initiation factor 5A (eIF5A) in dengue 2 virus-infected mosquito cells

Virology Journal ◽

10.1186/1743-422x-7-214 ◽

2010 ◽

Vol 7 (1) ◽

pp. 214 ◽

Cited By ~ 12

Author(s):

Yu-Tzu Shih ◽

Chao-Fu Yang ◽

Wei-June Chen

Keyword(s):

Translation Initiation ◽

Translation Initiation Factor ◽

Initiation Factor ◽

Infected Mosquito ◽

Eukaryotic Translation Initiation Factor ◽

Eukaryotic Translation ◽

Mosquito Cells ◽

Eukaryotic Translation Initiation

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Monitoring Mitochondrial Function in Aedes albopictus C6/36 Cell Line during Dengue Virus Infection

Insects ◽

10.3390/insects12100934 ◽

2021 ◽

Vol 12 (10) ◽

pp. 934

Author(s):

María E. Santana-Román ◽

Paola Maycotte ◽

Salvador Uribe-Carvajal ◽

Cristina Uribe-Alvarez ◽

Nayeli Alvarado-Medina ◽

...

Keyword(s):

Dengue Virus ◽

Cell Line ◽

Aedes Albopictus ◽

Mitochondrial Function ◽

Mitochondrial Fission ◽

Hemorrhagic Fever ◽

Denv Infection ◽

Infected Mosquito ◽

Mosquito Cells ◽

Induced Changes

Aedes aegypti and Aedes albopictus mosquitoes are responsible for dengue virus (DENV) transmission in tropical and subtropical areas worldwide, where an estimated 3 billion people live at risk of DENV exposure. DENV-infected individuals show symptoms ranging from sub-clinical or mild to hemorrhagic fever. Infected mosquitoes do not show detectable signs of disease, even though the virus maintains a lifelong persistent infection. The interactions between viruses and host mitochondria are crucial for virus replication and pathogenicity. DENV infection in vertebrate cells modulates mitochondrial function and dynamics to facilitate viral proliferation. Here, we describe that DENV also regulates mitochondrial function and morphology in infected C6/36 mosquito cells (derived from Aedes albopictus). Our results showed that DENV infection increased ROS (reactive oxygen species) production, modulated mitochondrial transmembrane potential and induced changes in mitochondrial respiration. Furthermore, we offer the first evidence that DENV causes translocation of mitofusins to mitochondria in the C6/36 mosquito cell line. Another protein Drp-1 (Dynamin-related protein 1) did not localize to mitochondria in DENV-infected cells. This observation therefore ruled out the possibility that the abovementioned alterations in mitochondrial function are associated with mitochondrial fission. In summary, this report provides some key insights into the virus–mitochondria crosstalk in DENV infected mosquito cells.

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Mosquito metabolomics reveal that dengue virus replication requires phospholipid reconfiguration via the remodeling cycle

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2015095117 ◽

2020 ◽

Vol 117 (44) ◽

pp. 27627-27636

Author(s):

Thomas Vial ◽

Wei-Lian Tan ◽

Eric Deharo ◽

Dorothée Missé ◽

Guillaume Marti ◽

...

Keyword(s):

Dengue Virus ◽

Viral Genome ◽

De Novo ◽

Genome Replication ◽

Mosquito Vector ◽

Phospholipid Biosynthesis ◽

Replication Complex ◽

Mosquito Cells ◽

Viral Genome Replication ◽

Blood Meals

Dengue virus (DENV) subdues cell membranes for its cellular cycle by reconfiguring phospholipids in humans and mosquitoes. Here, we determined how and why DENV reconfigures phospholipids in the mosquito vector. By inhibiting and activating the de novo phospholipid biosynthesis, we demonstrated the antiviral impact of de novo–produced phospholipids. In line with the virus hijacking lipids for its benefit, metabolomics analyses indicated that DENV actively inhibited the de novo phospholipid pathway and instead triggered phospholipid remodeling. We demonstrated the early induction of remodeling during infection by using isotope tracing in mosquito cells. We then confirmed in mosquitoes the antiviral impact of de novo phospholipids by supplementing infectious blood meals with a de novo phospholipid precursor. Eventually, we determined that phospholipid reconfiguration was required for viral genome replication but not for the other steps of the virus cellular cycle. Overall, we now propose that DENV reconfigures phospholipids through the remodeling cycle to modify the endomembrane and facilitate formation of the replication complex. Furthermore, our study identified de novo phospholipid precursor as a blood determinant of DENV human-to-mosquito transmission.

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Rapid selection against arbovirus-induced apoptosis during infection of a mosquito vector

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1424469112 ◽

2015 ◽

Vol 112 (10) ◽

pp. E1152-E1161 ◽

Cited By ~ 34

Author(s):

Katelyn O’Neill ◽

Bradley J. S. C. Olson ◽

Ning Huang ◽

Dave Unis ◽

Rollie J. Clem

Keyword(s):

Sindbis Virus ◽

Vector Competence ◽

Mosquito Species ◽

Infected Mosquito ◽

Nucleotide Sequencing ◽

Mosquito Vector ◽

Antiviral Defense ◽

Mosquito Cells ◽

Arbovirus Infection ◽

Induced Apoptosis

Millions of people are infected each year by arboviruses (arthropod-borne viruses) such as chikungunya, dengue, and West Nile viruses, yet for reasons that are largely unknown, only a relatively small number of mosquito species are able to transmit arboviruses. Understanding the complex factors that determine vector competence could facilitate strategies for controlling arbovirus infections. Apoptosis is a potential antiviral defense response that has been shown to be important in other virus–host systems. However, apoptosis is rarely seen in arbovirus-infected mosquito cells, raising questions about its importance as an antiviral defense in mosquitoes. We tested the effect of stimulating apoptosis during arbovirus infection by infectingAedes aegyptimosquitoes with a Sindbis virus (SINV) clone called MRE/Rpr, in which the MRE-16 strain of SINV was engineered to express the proapoptotic genereaperfromDrosophila. MRE/Rpr exhibited an impaired infection phenotype that included delayed midgut infection, delayed virus replication, and reduced virus accumulation in saliva. Nucleotide sequencing of thereaperinsert in virus populations isolated from individual mosquitoes revealed evidence of rapid and strong selection against maintenance of Reaper expression in MRE/Rpr-infected mosquitoes. The impaired phenotype of MRE/Rpr, coupled with the observed negative selection against Reaper expression, indicates that apoptosis is a powerful defense against arbovirus infection in mosquitoes and suggests that arboviruses have evolved mechanisms to avoid stimulating apoptosis in mosquitoes that serve as vectors.

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Comparative proteomics reveals that YK51, a 4-Hydroxypandurantin-A analogue, downregulates the expression of proteins associated with dengue virus infection

PeerJ ◽

10.7717/peerj.3939 ◽

2018 ◽

Vol 5 ◽

pp. e3939 ◽

Cited By ~ 4

Author(s):

Wei-Lian Tan ◽

Yean Kee Lee ◽

Yen Fong Ho ◽

Rohana Yusof ◽

Noorsaadah Abdul Rahman ◽

...

Keyword(s):

Virus Infection ◽

Dengue Virus ◽

Viral Infection ◽

Dengue Infection ◽

Functional Characterization ◽

Denv Infection ◽

Comparative Proteomics ◽

Therapeutic Drug ◽

Synthetic Analogue ◽

Down Regulation

Dengue is endemic throughout tropical and subtropical regions of the world. Currently, there is no clinically approved therapeutic drug available for this acute viral infection. Although the first dengue vaccine Dengvaxia has been approved for use in certain countries, it is limited to those without a previous dengue infection while the safety and efficacy of the vaccine in those elderly and younger children still need to be identified. Therefore, it is becoming increasingly important to develop therapeutics/drugs to combat dengue virus (DENV) infection. YK51 is a synthetic analogue of 4-Hydroxypandurantin A (a compound found in the crude extract of the rhizomes of Boesenbergia rotunda) that has been extensively studied by our research group. It has been shown to possess outstanding antiviral activity due to its inhibitory activity against NS2B/NS3 DENV2 protease. However, it is not known how YK51 affects the proteome of DENV infected cells. Therefore, we performed a comparative proteomics analysis to identify changes in protein expression in DENV infected HepG2 cells treated with YK51. Classical two-dimensional gel electrophoresis followed by protein identification using tandem mass spectrometry was employed in this study. Thirty proteins were found to be down-regulated with YK51 treatment. In silico analysis predicted that the down-regulation of eight of these proteins may inhibit viral infection. Our results suggested that apart from inhibiting the NS2B/NS3 DENV2 protease, YK51 may also be causing the down-regulation of a number of proteins that may be responsible in, and/or essential to virus infection. However, functional characterization of these proteins will be necessary before we can conclusively determine their roles in DENV infection.

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The Dengue Virus Nonstructural Protein 1 (NS1) Is Secreted from Mosquito Cells in Association with the Intracellular Cholesterol Transporter Chaperone Caveolin Complex

Journal of Virology ◽

10.1128/jvi.01985-18 ◽

2018 ◽

Vol 93 (4) ◽

Cited By ~ 5

Author(s):

Romel Rosales Ramirez ◽

Juan E. Ludert

Keyword(s):

Dengue Virus ◽

Nonstructural Protein ◽

Cell Interactions ◽

Vertebrate Host ◽

Mosquito Vector ◽

Caveolin 1 ◽

Nonstructural Protein 1 ◽

Virion Release ◽

Mosquito Cells ◽

Infected Cells

ABSTRACTDengue virus (DENV) is a mosquito-borne virus of the familyFlaviviridae. The RNA viral genome encodes three structural and seven nonstructural proteins. Nonstructural protein 1 (NS1) is a multifunctional protein actively secreted in vertebrate and mosquito cells during infection. In mosquito cells, NS1 is secreted in a caveolin-1-dependent manner by an unconventional route. The caveolin chaperone complex (CCC) is a cytoplasmic complex formed by caveolin-1 and the chaperones FKBP52, Cy40, and CyA and is responsible for the cholesterol traffic inside the cell. In this work, we demonstrate that in mosquito cells, but not in vertebrate cells, NS1 associates with and relies on the CCC for secretion. Treatment of mosquito cells with classic secretion inhibitors, such as brefeldin A, Golgicide A, and Fli-06, showed no effect on NS1 secretion but significant reductions in recombinant luciferase secretion and virion release. Silencing the expression of CAV-1 or FKBP52 with short interfering RNAs or the inhibition of CyA by cyclosporine resulted in significant decrease in NS1 secretion, again without affecting virion release. Colocalization, coimmunoprecipitation, and proximity ligation assays indicated that NS1 colocalizes and interacts with all proteins of the CCC. In addition, CAV-1 and FKBP52 expression was found augmented in DENV-infected cells. Results obtained with Zika virus-infected cells suggest that in mosquito cells, ZIKV NS1 follows the same secretory pathway as that observed for DENV NS1. These results uncover important differences in the dengue virus-cell interactions between the vertebrate host and the mosquito vector as well as novel functions for the chaperone caveolin complex.IMPORTANCEThe dengue virus protein NS1 is secreted efficiently from both infected vertebrate and mosquito cells. Previously, our group reported that NS1 secretion in mosquito cells follows an unconventional secretion pathway dependent on caveolin-1. In this work, we demonstrate that in mosquito cells, but not in vertebrate cells, NS1 secretion takes place in association with the chaperone caveolin complex, a complex formed by caveolin-1 and the chaperones FKBP52, CyA, and Cy40, which are in charge of cholesterol transport inside the cell. Results obtained with ZIKV-infected mosquito cells suggest that ZIKV NS1 is released following an unconventional secretory route in association with the chaperone caveolin complex. These results uncover important differences in the virus-cell interactions between the vertebrate host and the mosquito vector, as well as novel functions for the chaperone caveolin complex. Moreover, manipulation of the NS1 secretory route may prove a valuable strategy to combat these two mosquito-borne diseases.

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Ultrastructural Characterization and Three-Dimensional Architecture of Replication Sites in Dengue Virus-Infected Mosquito Cells

Journal of Virology ◽

10.1128/jvi.00118-14 ◽

2014 ◽

Vol 88 (9) ◽

pp. 4687-4697 ◽

Cited By ~ 113

Author(s):

J. Junjhon ◽

J. G. Pennington ◽

T. J. Edwards ◽

R. Perera ◽

J. Lanman ◽

...

Keyword(s):

Dengue Virus ◽

Three Dimensional ◽

Infected Mosquito ◽

Mosquito Cells ◽

Ultrastructural Characterization ◽

Replication Sites

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Aedes aegypti SNAP and a calcium transporter ATPase influence dengue virus dissemination

10.1101/2020.07.10.198226 ◽

2020 ◽

Author(s):

Alejandro Marin-Lopez ◽

Junjun Jiang ◽

Yuchen Wang ◽

Yongguo Cao ◽

Tyler MacNeil ◽

...

Keyword(s):

Salivary Gland ◽

Aedes Aegypti ◽

Dengue Virus ◽

Denv Infection ◽

Infected Mosquito ◽

Viral Dissemination ◽

Protein Components ◽

Calcium Transporter

AbstractDengue virus (DENV) is a flavivirus that causes marked human morbidity and mortality worldwide, being transmitted to humans by Aedes aegypti mosquitoes. Habitat expansion of Aedes, mainly due to climate change and increasing overlap between urban and wild habitats, places nearly half of the world’s population at risk for DENV infection. After a bloodmeal from a DENV-infected host, the virus enters the mosquito midgut. Next, the virus migrates to, and replicates in, other tissues, like salivary glands. Successful viral transmission occurs when the infected mosquito takes another blood meal on a susceptible host and DENV is released from the salivary gland via saliva into the skin. During viral dissemination in the mosquito and transmission to a new mammalian host, DENV interacts with a variety of vector proteins, which are uniquely important during each phase of the viral cycle. Our study focuses on the interaction between DENV particles and protein components in the A. aegypti vector. We performed a mass spectrometry assay where we identified a set of A aegypti salivary gland proteins which potentially interact with the DENV virion. Using dsRNA to silence gene expression, we analyzed the role of these proteins in viral infectivity. Two of these candidates, a synaptosomal-associated protein (AeSNAP) and a calcium transporter ATPase (ATPase) appear to play a role in viral replication both in vitro and in vivo. These findings suggest that AeSNAP plays a protective role during DENV infection of mosquitoes and that ATPase protein is required for DENV during amplification within the vector.ImportanceAedes aegypti mosquitoes are the major vector of different flaviviruses that cause human diseases, including dengue virus. There is a great need for better therapeutics and preventive vaccines against flaviviruses. Flaviviruses create complex virus-host and virus-vector interactions. The interactions between viral particles and protein components in the vector is not completely understood. In this work we characterize how two mosquito proteins, “AeSNAP” and “ATPase”, influence DENV viral dissemination within A. aegypti, using both in vitro and in vivo models. These results elucidate anti-vector measures that may be potentially be used to control dengue virus spread in the mosquito vector.

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Secretion of Nonstructural Protein 1 of Dengue Virus from Infected Mosquito Cells: Facts and Speculations

Journal of Virology ◽

10.1128/jvi.00275-18 ◽

2018 ◽

Vol 92 (14) ◽

Cited By ~ 4

Author(s):

Ana C. Alcalá ◽

Laura A. Palomares ◽

Juan E. Ludert

Keyword(s):

Dengue Virus ◽

Nonstructural Protein ◽

Infected Mosquito ◽

Nonstructural Protein 1 ◽

Mosquito Cells

ABSTRACTDengue virus nonstructural protein 1 (NS1) is a multifunctional glycoprotein. For decades, the notion in the field was that NS1 is secreted exclusively from vertebrate cells and not from mosquito cells. However, recent evidence shows that mosquito cells also secrete NS1 efficiently. In this review, we discuss the evidence for secretion of NS1 of dengue virus, and of other flaviviruses, from mosquito cells, differences between NS1 secreted from mosquito and NS1 secreted from vertebrate cells, and possible roles of soluble NS1 in the insect flavivirus vector.

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