Host Cell Transcriptome Profile During Wild-type and Attenuated Dengue Virus Infection

PLoS Negl Trop Dis. 2013;7(3):e2107.

October M. Sessions§1, Ying Tan§1, Kenneth C. Goh1, Yujing Liu2,3, Patrick Tan4, Steve Rozen2, Eng Eong Ooi1*

 

1Program in Emerging Infectious Disease, Duke-NUS Graduate Medical School, Singapore

2Centre for Computational Biology, Duke-NUS Graduate Medical School, Singapore

3Computational Systems Biology, Singapore-MIT Alliance, National University of Singapore, Singapore

4Program in Cancer and Stem Cell Biology, Duke-NUS Graduate Medical School, Singapore

§Co-first authors

*Corresponding author

 

Abstract

Dengue viruses 1-4 (DENV1-4) rely heavily on the host cell machinery to complete their life cycle, while at the same time evade the host response that could restrict their replication efficiency.  These requirements may account for much of the broad gene-level changes to the host transcriptome upon DENV infection.  However, host gene function is also regulated through transcriptional start site (TSS) selection and post-transcriptional modification to the RNA that give rise to multiple gene isoforms.  The roles these processes play in the host response to dengue infection have not been explored.  In the present study, we utilized RNA sequencing (RNAseq) to identify novel transcript variations in response to infection with both a pathogenic strain of DENV1 and its attenuated derivative.  RNAseq provides the information necessary to distinguish the various isoforms produced from a single gene and their splice variants.  Our data indicate that there is an extensive amount of previously uncharacterized TSS and post-transcriptional modifications to host RNA over a wide range of pathways and host functions in response to DENV infection.  Many of the differentially expressed genes identified in this study have previously been shown to be required for flavivirus propagation and/or interact with DENV gene products.  We also show here that the human transcriptome response to an infection by wild-type DENV or its attenuated derivative differs significantly.  This differential response to wild-type and attenuated DENV infection suggests that alternative processing events may be part of a previously uncharacterized innate immune response to viral infection that is in large part evaded by wild-type DENV.

PMID: 23516652

 

Summary

Dengue is the most common vector-borne viral disease globally, affecting an estimated 400 million people each year.  The continued absence of an effective therapy stems from an incomplete understanding of disease pathogenesis, of which the host response to infection is thought to play a central role.  All previous studies that examined the host response to dengue virus infection have relied on the microarray technology.  While those studies have described the changes in gene expression with dengue virus infection, they have not been able to provide any information on transcriptional start site and post-transcriptional variations of the host RNA. These variations lead to the production of gene isoforms that could have a profound effect on host response.  Understanding RNA processing during dengue virus infection could provide new insights into dengue pathogenesis and identify new targets for therapeutic intervention.

In an attempt to create a live-attenuated vaccine for dengue virus serotype 1, the clinical isolate DENV1-16007 was serially passaged in primary dog kidney cells for 13 generations to produce the DENV1-PDK13 strain.  DENV1-PDK13 was taken through phase II clinical trials and was shown to be both immunogenic and minimally reactogenic, the hallmarks of a good live-attenuated vaccine.  Although the final vaccine formulation was unsuccessful due to incomplete attenuation of the dengue serotype 3 constituent, the 16007/PDK13 pair provided us with an excellent opportunity to investigate the transcriptional differences in the host response to infection by two highly related, but phenotypically different dengue viruses and possibly gain a mechanistic insight into disease pathogenesis.  To study these differences with an unprecedented amount of detail, we used deep sequencing to interrogate the transcriptional changes down to the level of transcriptional start site selection, isoform variation and splicing.  These analyses allowed us to draw to two major conclusions:

  1. There is an extensive amount of previously uncharacterized transcriptional start site and post-transcriptional variation in host RNA over a wide range of pathways and host functions in response to DENV infection.
  2. The human transcriptomic response to an infection by wild-type DENV or differs significantly to its attenuated derivative, which suggests that alternative processing events may be part of a previously uncharacterized innate immune response to viral infection that is in large part evaded by wild-type DENV.

We are cognizant of the reality that some portion of the hits identified in large-scale genomics studies such as this one may be platform or cell line specific and may not extend to other model systems or in vivo observations.  In order provide a useful resource to the research community interested in dengue pathogenesis, we therefore sought to frame the results of this study into the broader context of the existing body of knowledge.  To do this, we conducted a cross-platform integrative analysis of our observations with previous studies that identified genes that are either differentially regulated or are thought to serve as host factors for flavivirus replication (Figure 1).  When taken collectively, the data demonstrates that specific pathways are differentially regulated at the transcriptional and post-transcriptional levels, which could serve as signatures of viral attenuation for vaccine development.

Eng Eong-1

Figure 1 – Making sense of large genomic data sets. We envisage that the data from this analysis could serve as a catalyst for hypothesis-driven studies on the role of these factors and their respective isoforms in dengue infection.

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