scholarly journals Subgenomic flavivirus RNA binds the mosquito DEAD/H-box helicase ME31B and determines Zika virus transmission by Aedes aegypti

2019 ◽  
Vol 116 (38) ◽  
pp. 19136-19144 ◽  
Author(s):  
Giel P. Göertz ◽  
Joyce W. M. van Bree ◽  
Anwar Hiralal ◽  
Bas M. Fernhout ◽  
Carmen Steffens ◽  
...  
Keyword(s):  
Aedes Aegypti ◽  
Zika Virus ◽  
Affinity Purification ◽  
Mechanistic Model ◽  
Virus Transmission ◽  
Protein Translation ◽  
Mass Spectrometry Analysis ◽  
Small Interfering Rnas ◽  
Spectrometry Analysis ◽  
Mosquito Infection

Zika virus (ZIKV) is an arthropod-borne flavivirus predominantly transmitted by Aedes aegypti mosquitoes and poses a global human health threat. All flaviviruses, including those that exclusively replicate in mosquitoes, produce a highly abundant, noncoding subgenomic flavivirus RNA (sfRNA) in infected cells, which implies an important function of sfRNA during mosquito infection. Currently, the role of sfRNA in flavivirus transmission by mosquitoes is not well understood. Here, we demonstrate that an sfRNA-deficient ZIKV (ZIKVΔSF1) replicates similar to wild-type ZIKV in mosquito cell culture but is severely attenuated in transmission by Ae. aegypti after an infectious blood meal, with 5% saliva-positive mosquitoes for ZIKVΔSF1 vs. 31% for ZIKV. Furthermore, viral titers in the mosquito saliva were lower for ZIKVΔSF1 as compared to ZIKV. Comparison of mosquito infection via infectious blood meals and intrathoracic injections showed that sfRNA is important for ZIKV to overcome the mosquito midgut barrier and to promote virus accumulation in the saliva. Next-generation sequencing of infected mosquitoes showed that viral small-interfering RNAs were elevated upon ZIKVΔSF1 as compared to ZIKV infection. RNA-affinity purification followed by mass spectrometry analysis uncovered that sfRNA specifically interacts with a specific set of Ae. aegypti proteins that are normally associated with RNA turnover and protein translation. The DEAD/H-box helicase ME31B showed the highest affinity for sfRNA and displayed antiviral activity against ZIKV in Ae. aegypti cells. Based on these results, we present a mechanistic model in which sfRNA sequesters ME31B to promote flavivirus replication and virion production to facilitate transmission by mosquitoes.

scholarly journals Re-Discovery of Giardiavirus: Genomic and Functional Analysis of Viruses from Giardia duodenalis Isolates

Biomedicines ◽  
2021 ◽  
Vol 9 (6) ◽  
pp. 654
Author(s):  
Gianluca Marucci ◽  
Ilaria Zullino ◽  
Lucia Bertuccini ◽  
Serena Camerini ◽  
Serena Cecchetti ◽  
...  
Keyword(s):  
High Throughput Sequencing ◽  
Diarrheal Disease ◽  
Current Knowledge ◽  
Biological Significance ◽  
Giardia Duodenalis ◽  
Protozoan Parasite ◽  
Protein Translation ◽  
Mass Spectrometry Analysis ◽  
Animal Origin ◽  
Spectrometry Analysis

Giardiasis, caused by the protozoan parasite Giardia duodenalis, is an intestinal diarrheal disease affecting almost one billion people worldwide. A small endosymbiotic dsRNA viruses, G. lamblia virus (GLV), genus Giardiavirus, family Totiviridae, might inhabit human and animal isolates of G. duodenalis. Three GLV genomes have been sequenced so far, and only one was intensively studied; moreover, a positive correlation between GLV and parasite virulence is yet to be proved. To understand the biological significance of GLV infection in Giardia, the characterization of several GLV strains from naturally infected G. duodenalis isolates is necessary. Here we report high-throughput sequencing of four GLVs strains, from Giardia isolates of human and animal origin. We also report on a new, unclassified viral sequence (designed GdRV-2), unrelated to Giardiavirus, encoding and expressing for a single large protein with an RdRp domain homologous to Totiviridae and Botybirnaviridae. The result of our sequencing and proteomic analyses challenge the current knowledge on GLV and strongly suggest that viral capsid protein translation unusually starts with a proline and that translation of the RNA-dependent RNA polymerase (RdRp) occurs via a +1/−2 ribosomal frameshift mechanism. Nucleotide polymorphism, confirmed by mass-spectrometry analysis, was also observed among and between GLV strains. Phylogenetic analysis indicated the occurrence of at least two GLV subtypes which display different phenotypes and transmissibility in experimental infections of a GLV naïve Giardia isolate.

scholarly journals Sequential Infection of Aedes aegypti Mosquitoes with Chikungunya Virus and Zika Virus Enhances Early Zika Virus Transmission

Insects ◽  
2018 ◽  
Vol 9 (4) ◽  
pp. 177 ◽  
Author(s):  
Tereza Magalhaes ◽  
Alexis Robison ◽  
Michael Young ◽  
William Black ◽  
Brian Foy ◽  
...  
Keyword(s):  
Aedes Aegypti ◽  
Blood Meal ◽  
Zika Virus ◽  
Chikungunya Virus ◽  
Vector Competence ◽  
Virus Transmission ◽  
Mosquito Vector ◽  
Molecular Aspects ◽  
Virus Interactions ◽  
Sequential Infection

In urban settings, chikungunya, Zika, and dengue viruses are transmitted by Aedes aegypti mosquitoes. Since these viruses co-circulate in several regions, coinfection in humans and vectors may occur, and human coinfections have been frequently reported. Yet, little is known about the molecular aspects of virus interactions within hosts and how they contribute to arbovirus transmission dynamics. We have previously shown that Aedes aegypti exposed to chikungunya and Zika viruses in the same blood meal can become coinfected and transmit both viruses simultaneously. However, mosquitoes may also become coinfected by multiple, sequential feeds on single infected hosts. Therefore, we tested whether sequential infection with chikungunya and Zika viruses impacts mosquito vector competence. We exposed Ae. aegypti mosquitoes first to one virus and 7 days later to the other virus and compared infection, dissemination, and transmission rates between sequentially and single infected groups. We found that coinfection rates were high after sequential exposure and that mosquitoes were able to co-transmit both viruses. Surprisingly, chikungunya virus coinfection enhanced Zika virus transmission 7 days after the second blood meal. Our data demonstrate heterologous arbovirus synergism within mosquitoes, by unknown mechanisms, leading to enhancement of transmission under certain conditions.

scholarly journals Identification of sulfenylated cysteines in Arabidopsis thaliana proteins using a disulfide-linked peptide reporter

2020 ◽  
Author(s):  
Bo Wei ◽  
Patrick Willems ◽  
Jingjing Huang ◽  
Caiping Tian ◽  
Jing Yang ◽  
...  
Keyword(s):  
Arabidopsis Thaliana ◽  
Protein Interactions ◽  
Affinity Purification ◽  
Mass Spectrometry Analysis ◽  
Technological Advancement ◽  
Amino Acid Residues ◽  
Cysteine Oxidation ◽  
Spectrometry Analysis ◽  
Mixed Disulfide ◽  
And Function

ABSTRACTIn proteins, hydrogen peroxide (H2O2) reacts with redox-sensitive cysteines to form cysteine sulfenic acid, also known as S-sulfenylation. These cysteine oxidation events can steer diverse cellular processes by altering protein interactions, trafficking, conformation, and function. Previously, we had identified S-sulfenylated proteins by using a tagged proteinaceous probe based on the yeast AP-1–like (Yap1) transcription factor that specifically reacts with sulfenic acids and traps them through a mixed disulfide bond. However, the identity of the S-sulfenylated amino acid residues remained enigmatic. Here, we present a technological advancement to identify in situ sulfenylated cysteines directly by means of the transgenic Yap1 probe. In Arabidopsis thaliana cells, after an initial affinity purification and a tryptic digestion, we further enriched the mixed disulfide-linked peptides with an antibody targeting the YAP1C-derived peptide (C598SEIWDR) that entails the redox-active cysteine. Subsequent mass spectrometry analysis with pLink 2 identified 1,745 YAP1C cross-linked peptides, indicating sulfenylated cysteines in over 1,000 proteins. Approximately 55% of these YAP1C-linked cysteines had previously been reported as redox-sensitive cysteines (S-sulfenylation, S-nitrosylation, and reversibly oxidized cysteines). The presented methodology provides a noninvasive approach to identify sulfenylated cysteines in any species that can be genetically modified.

scholarly journals siRNA Design to Silence the 3′UTR Region of Zika Virus

2020 ◽  
Vol 2020 ◽  
pp. 1-8
Author(s):  
María Perez-Mendez ◽  
Paola Zárate-Segura ◽  
Juan Salas-Benito ◽  
Fernando Bastida-González
Keyword(s):  
Public Health ◽  
Blood Transfusion ◽  
Aedes Aegypti ◽  
Cell Line ◽  
Birth Defects ◽  
Zika Virus ◽  
In Silico ◽  
Small Interfering Rnas ◽  
Medical Problems ◽  
Sirna Design

The disease caused by the Zika virus (ZIKV) has positioned itself as one of the main public health problems in Mexico. One of the main reasons is it causes microcephaly and other birth defects. The transmission of ZIKV is through Aedes aegypti and Ae. albopictus mosquitoes, which are found in a larger space of the national territory. In addition, it can also be transmitted via blood transfusion, sexual relations, and maternal-fetal route. So far, there are no vaccines or specific treatments to deal with this infection. Currently, some new therapeutics such as small interfering RNAs (siRNAs) are able to regulate or suppress transcription in viruses. Therefore, in this project, an in silico siRNA was designed for the 3′UTR region of ZIKV via bioinformatics tools. The designed siRNA was synthesized and transfected into the C6/36 cell line, previously infected with ZIKV in order to assess the ability of the siRNA to inhibit viral replication. The designed siRNA was able to inhibit significantly (p<0.05) ZIKV replication; this siRNA could be considered a potential therapeutic towards the disease that causes ZIKV and the medical problems generated.

scholarly journals ProbingN-glycoprotein microheterogeneity by lectin affinity purification-mass spectrometry analysis

2019 ◽  
Vol 10 (19) ◽  
pp. 5146-5155 ◽  
Author(s):  
Di Wu ◽  
Jingwen Li ◽  
Weston B. Struwe ◽  
Carol V. Robinson
Keyword(s):  
Mass Spectrometry ◽  
Protein Level ◽  
Affinity Purification ◽  
Mass Spectrometry Analysis ◽  
Intact Protein ◽  
Spectrometry Analysis ◽  
Lectin Affinity ◽  
Affinity Purification Mass Spectrometry

A lectin affinity purification-mass spectrometry approach to characterize lectin-reactive glycoproteoforms and elucidate lectin specificities at the intact protein level.

scholarly journals Wolbachia strain w AlbA blocks Zika virus transmission in Aedes aegypti

2019 ◽  
Vol 34 (1) ◽  
pp. 116-119 ◽  
Author(s):  
T. Chouin‐Carneiro ◽  
T. H. Ant ◽  
C. Herd ◽  
F. Louis ◽  
A. B. Failloux ◽  
...  
Keyword(s):  
Aedes Aegypti ◽  
Zika Virus ◽  
Virus Transmission ◽  
Wolbachia Strain

scholarly journals SAMHD1 Enhances Chikungunya and Zika Virus Replication in Human Skin Fibroblasts

Proceedings ◽  
2020 ◽  
Vol 50 (1) ◽  
pp. 81
Author(s):  
Sineewanlaya Wichit
Keyword(s):  
Human Skin ◽  
Zika Virus ◽  
Antiviral Treatment ◽  
Skin Fibroblasts ◽  
Mass Spectrometry Analysis ◽  
Strong Increase ◽  
Human Skin Fibroblasts ◽  
Entry Site ◽  
Stable Isotope Labelling ◽  
Spectrometry Analysis

Chikungunya virus (CHIKV) and Zika virus (ZIKV) are emerging arboviruses that pose a worldwide threat to human health. Currently, neither vaccine nor antiviral treatment to control their infections is available. As the skin is a major viral entry site for arboviruses in the human host, we determined the global proteomic profile of CHIKV and ZIKV infections in human skin fibroblasts using stable isotope labelling by amino acids in cell culture (SILAC)-based mass spectrometry analysis. We show that the expressions of the interferon-stimulated proteins MX1, IFIT1, IFIT3 and ISG15, as well as expressions of defense response proteins DDX58, STAT1, OAS3, EIF2AK2, and SAMHD1 were significantly upregulated in these cells upon infection with either virus. Exogenous expression of IFITs proteins markedly inhibited CHIKV and ZIKV replication which, accordingly, was restored following the abrogation of IFIT1 or IFIT3. Overexpression of SAMHD1 in cutaneous cells or pretreatment of cells with the virus-like particles containing SAMHD1 restriction factor Vpx resulted in a strong increase or inhibition, respectively, in both CHIKV and ZIKV replication. Moreover, silencing of SAMHD1 by specific SAMHD1-siRNA resulted in a marked decrease in viral RNA levels. Together, these results suggest that IFITs are involved in the restriction of replication of CHIKV and ZIKV and provide, as yet unreported, evidence for a proviral role of SAMHD1 in arbovirus infection of human skin cells.

scholarly journals Identification of Novel Surface Proteins of Anaplasma phagocytophilum by Affinity Purification and Proteomics

2007 ◽  
Vol 189 (21) ◽  
pp. 7819-7828 ◽  
Author(s):  
Yan Ge ◽  
Yasuko Rikihisa
Keyword(s):  
Anaplasma Phagocytophilum ◽  
Affinity Purification ◽  
The United States ◽  
Capillary Liquid Chromatography ◽  
Etiologic Agent ◽  
Surface Proteins ◽  
Mass Spectrometry Analysis ◽  
Spectrometry Analysis ◽  
Major Surface

ABSTRACT Anaplasma phagocytophilum is the etiologic agent of human granulocytic anaplasmosis (HGA), one of the major tick-borne zoonoses in the United States. The surface of A. phagocytophilum plays a crucial role in subverting the hostile host cell environment. However, except for the P44/Msp2 outer membrane protein family, the surface components of A. phagocytophilum are largely unknown. To identify the major surface proteins of A. phagocytophilum, a membrane-impermeable, cleavable biotin reagent, sulfosuccinimidyl-2-[biotinamido]ethyl-1,3-dithiopropionate (Sulfo-NHS-SS-Biotin), was used to label intact bacteria. The biotinylated bacterial surface proteins were isolated by streptavidin agarose affinity purification and then separated by electrophoresis, followed by capillary liquid chromatography-nanospray tandem mass spectrometry analysis. Among the major proteins captured by affinity purification were five A. phagocytophilum proteins, Omp85, hypothetical proteins APH_0404 (designated Asp62) and APH_0405 (designated Asp55), P44 family proteins, and Omp-1A. The surface exposure of Asp62 and Asp55 was verified by immunofluorescence microscopy. Recombinant Asp62 and Asp55 proteins were recognized by an HGA patient serum. Anti-Asp62 and anti-Asp55 peptide sera partially neutralized A. phagocytophilum infection of HL-60 cells in vitro. We found that the Asp62 and Asp55 genes were cotranscribed and conserved among members of the family Anaplasmataceae. With the exception of P44-18, all of the proteins were newly revealed major surface-exposed proteins whose study should facilitate understanding the interaction between A. phagocytophilum and the host. These proteins may serve as targets for development of chemotherapy, diagnostics, and vaccines.

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