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Wolbachia Blocks Viral Genome Replication Early in Infection without a Transcriptional Response by the Endosymbiont or Host Small RNA Pathways.

Rainey SM, Martinez J, McFarlane M, Juneja P, Sarkies P, Lulla A, Schnettler E, Varjak M, Merits A, Miska EA, Jiggins FM, Kohl A - PLoS Pathog. (2016)

Bottom Line: Furthermore, it causes a far greater reduction in the expression of proteins from the 3' open reading frame than the 5' non-structural protein open reading frame, indicating that it is blocking the replication of viral RNA.Further to this separation of the replicase proteins and viral RNA in transreplication assays shows that uncoupling of viral RNA and replicase proteins does not overcome Wolbachia's antiviral activity.Host microRNAs (miRNAs) have been implicated in protection, but again we found that host cell miRNA expression was unaffected by the bacterium and neither do our findings suggest any involvement of the antiviral siRNA pathway.

View Article: PubMed Central - PubMed

Affiliation: MRC-University of Glasgow Centre for Virus Research, Glasgow, Scotland, United Kingdom.

ABSTRACT
The intracellular endosymbiotic bacterium Wolbachia can protect insects against viral infection, and is being introduced into mosquito populations in the wild to block the transmission of arboviruses that infect humans and are a major public health concern. To investigate the mechanisms underlying this antiviral protection, we have developed a new model system combining Wolbachia-infected Drosophila melanogaster cell culture with the model mosquito-borne Semliki Forest virus (SFV; Togaviridae, Alphavirus). Wolbachia provides strong antiviral protection rapidly after infection, suggesting that an early stage post-infection is being blocked. Wolbachia does appear to have major effects on events distinct from entry, assembly or exit as it inhibits the replication of an SFV replicon transfected into the cells. Furthermore, it causes a far greater reduction in the expression of proteins from the 3' open reading frame than the 5' non-structural protein open reading frame, indicating that it is blocking the replication of viral RNA. Further to this separation of the replicase proteins and viral RNA in transreplication assays shows that uncoupling of viral RNA and replicase proteins does not overcome Wolbachia's antiviral activity. This further suggests that replicative processes are disrupted, such as translation or replication, by Wolbachia infection. This may occur by Wolbachia mounting an active antiviral response, but the virus did not cause any transcriptional response by the bacterium, suggesting that this is not the case. Host microRNAs (miRNAs) have been implicated in protection, but again we found that host cell miRNA expression was unaffected by the bacterium and neither do our findings suggest any involvement of the antiviral siRNA pathway. We conclude that Wolbachia may directly interfere with early events in virus replication such as translation of incoming viral RNA or RNA transcription, and this likely involves an intrinsic (as opposed to an induced) mechanism.

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The effect of Wolbachia on D. melanogaster miRNA expression in the presence or absence of virus.Heatmap showing the effects of Wolbachia and SFV4(3H)-RLuc virus on miRNA abundance. The mean of the No Virus-No Wolbachia treatment is set to zero. Only miRNAs that are significantly (Adjusted p value<0.1, Negative Binomial Test) differentially expressed in at least one treatment are shown. Jw18Wol replicate WV 3 was included in analysis; although it appeared to be an outlier as its inclusion did not alter the significance of the data. T = Jw18Free, W = Jw18Wol, V = SFV infection, O = no SFV infection and numbers represent replicate.
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ppat.1005536.g006: The effect of Wolbachia on D. melanogaster miRNA expression in the presence or absence of virus.Heatmap showing the effects of Wolbachia and SFV4(3H)-RLuc virus on miRNA abundance. The mean of the No Virus-No Wolbachia treatment is set to zero. Only miRNAs that are significantly (Adjusted p value<0.1, Negative Binomial Test) differentially expressed in at least one treatment are shown. Jw18Wol replicate WV 3 was included in analysis; although it appeared to be an outlier as its inclusion did not alter the significance of the data. T = Jw18Free, W = Jw18Wol, V = SFV infection, O = no SFV infection and numbers represent replicate.

Mentions: Previous studies suggested that Wolbachia affects the sensitivity of mosquito cells to viral infection by altering host miRNAs levels [31, 32]. Therefore, we tested whether Wolbachia alters the expression of known miRNAs in Jw18 cells. In the absence of virus, no miRNAs had significantly different expression levels in Jw18Wol and Jw18Free cells (Fig 6 and S3A Fig). It is likely therefore that any differences are not important to Wolbachia mediated protection as in our system there are no significantly altered miRNAs between non-infected Jw18Free and Jw18Wol cells.


Wolbachia Blocks Viral Genome Replication Early in Infection without a Transcriptional Response by the Endosymbiont or Host Small RNA Pathways.

Rainey SM, Martinez J, McFarlane M, Juneja P, Sarkies P, Lulla A, Schnettler E, Varjak M, Merits A, Miska EA, Jiggins FM, Kohl A - PLoS Pathog. (2016)

The effect of Wolbachia on D. melanogaster miRNA expression in the presence or absence of virus.Heatmap showing the effects of Wolbachia and SFV4(3H)-RLuc virus on miRNA abundance. The mean of the No Virus-No Wolbachia treatment is set to zero. Only miRNAs that are significantly (Adjusted p value<0.1, Negative Binomial Test) differentially expressed in at least one treatment are shown. Jw18Wol replicate WV 3 was included in analysis; although it appeared to be an outlier as its inclusion did not alter the significance of the data. T = Jw18Free, W = Jw18Wol, V = SFV infection, O = no SFV infection and numbers represent replicate.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4835223&req=5

ppat.1005536.g006: The effect of Wolbachia on D. melanogaster miRNA expression in the presence or absence of virus.Heatmap showing the effects of Wolbachia and SFV4(3H)-RLuc virus on miRNA abundance. The mean of the No Virus-No Wolbachia treatment is set to zero. Only miRNAs that are significantly (Adjusted p value<0.1, Negative Binomial Test) differentially expressed in at least one treatment are shown. Jw18Wol replicate WV 3 was included in analysis; although it appeared to be an outlier as its inclusion did not alter the significance of the data. T = Jw18Free, W = Jw18Wol, V = SFV infection, O = no SFV infection and numbers represent replicate.
Mentions: Previous studies suggested that Wolbachia affects the sensitivity of mosquito cells to viral infection by altering host miRNAs levels [31, 32]. Therefore, we tested whether Wolbachia alters the expression of known miRNAs in Jw18 cells. In the absence of virus, no miRNAs had significantly different expression levels in Jw18Wol and Jw18Free cells (Fig 6 and S3A Fig). It is likely therefore that any differences are not important to Wolbachia mediated protection as in our system there are no significantly altered miRNAs between non-infected Jw18Free and Jw18Wol cells.

Bottom Line: Furthermore, it causes a far greater reduction in the expression of proteins from the 3' open reading frame than the 5' non-structural protein open reading frame, indicating that it is blocking the replication of viral RNA.Further to this separation of the replicase proteins and viral RNA in transreplication assays shows that uncoupling of viral RNA and replicase proteins does not overcome Wolbachia's antiviral activity.Host microRNAs (miRNAs) have been implicated in protection, but again we found that host cell miRNA expression was unaffected by the bacterium and neither do our findings suggest any involvement of the antiviral siRNA pathway.

View Article: PubMed Central - PubMed

Affiliation: MRC-University of Glasgow Centre for Virus Research, Glasgow, Scotland, United Kingdom.

ABSTRACT
The intracellular endosymbiotic bacterium Wolbachia can protect insects against viral infection, and is being introduced into mosquito populations in the wild to block the transmission of arboviruses that infect humans and are a major public health concern. To investigate the mechanisms underlying this antiviral protection, we have developed a new model system combining Wolbachia-infected Drosophila melanogaster cell culture with the model mosquito-borne Semliki Forest virus (SFV; Togaviridae, Alphavirus). Wolbachia provides strong antiviral protection rapidly after infection, suggesting that an early stage post-infection is being blocked. Wolbachia does appear to have major effects on events distinct from entry, assembly or exit as it inhibits the replication of an SFV replicon transfected into the cells. Furthermore, it causes a far greater reduction in the expression of proteins from the 3' open reading frame than the 5' non-structural protein open reading frame, indicating that it is blocking the replication of viral RNA. Further to this separation of the replicase proteins and viral RNA in transreplication assays shows that uncoupling of viral RNA and replicase proteins does not overcome Wolbachia's antiviral activity. This further suggests that replicative processes are disrupted, such as translation or replication, by Wolbachia infection. This may occur by Wolbachia mounting an active antiviral response, but the virus did not cause any transcriptional response by the bacterium, suggesting that this is not the case. Host microRNAs (miRNAs) have been implicated in protection, but again we found that host cell miRNA expression was unaffected by the bacterium and neither do our findings suggest any involvement of the antiviral siRNA pathway. We conclude that Wolbachia may directly interfere with early events in virus replication such as translation of incoming viral RNA or RNA transcription, and this likely involves an intrinsic (as opposed to an induced) mechanism.

Show MeSH
Related in: MedlinePlus