Comparative Transcriptomics Highlights the Role of the Activator Protein 1 Transcription Factor in the Host Response to Ebolavirus

Abstract

Ebolavirus and Marburgvirus comprise two genera of negative-sense singlestranded RNA viruses that cause severe hemorrhagic fevers in humans. Despite considerable research efforts, the molecular events following Ebola virus (EBOV) infection are poorly understood. With the view of identifying host factors that underpin EBOV pathogenesis, we compared the transcriptomes of EBOV-infected human, pig, and bat kidney cells using a transcriptome sequencing (RNA-seq) approach. Despite a significant difference in viral transcription/replication between the cell lines, all cells responded to EBOV infection through a robust induction of extracellular growth factors. Furthermore, a significant upregulation of activator protein 1 (AP1) transcription factor complex members FOS and JUN was observed in permissive cell lines. Functional studies focusing on human cells showed that EBOV infection induces protein expression, phosphorylation, and nuclear accumulation of JUN and, to a lesser degree, FOS. Using a luciferase-based reporter, we show that EBOV infection induces AP1 transactivation activity within human cells at 48 and 72 h postinfection. Finally, we show that JUN knockdown decreases the expression of EBOV-induced host gene expression. Taken together, our study highlights the role of AP1 in promoting the host gene expression profile that defines EBOV pathogenesis. IMPORTANCE Many questions remain about the molecular events that underpin filovirus pathophysiology. The rational design of new intervention strategies, such as postexposure therapeutics, will be significantly enhanced through an in-depth understanding of these molecular events. We believe that new insights into the molecular pathogenesis of EBOV may be possible by examining the transcriptomic response of taxonomically diverse cell lines (derived from human, pig, and bat). We first identified the responsive pathways using an RNA-seq-based transcriptomics approach. Further functional and computational analysis focusing on human cells highlighted an important role for the AP1 transcription factor in mediating the transcriptional response to EBOV infection. Our study sheds new light on how host transcription factors respond to and promote the transcriptional landscape that follows viral infection.

Bat-borne virus diversity, spillover and emergence

“…Interferon-ɑ (IFNα), IFNβ and IFNɣ pathways vary in their level of activation between bat and human cells in response to viral infection [49][50][51][52] . Some of these studies have shown dampened immune responses in bats, whereas others have shown heightened responses to infection.…”

Utility of primary cells to examine NPC1 receptor expression in Mops condylurus, a potential Ebola virus reservoir

“…These results reflect the possible dramatic change of the characteristics of cell cultures within a very short time. Although the vast majority of cell culture experiments in filovirus research, including model systems for entry and replication, are performed with immortalized cell lines [36,[41][42][43][44][45][46][47][48][49], the limitations of these model systems should be taken into consideration. We have also demonstrated the importance of the use of cells in low passage numbers not taken into consideration in many studies.…”

“…The examination of EBOV entry and infection processes in bat cells have predominantly been performed in immortalized cell lines [36,[41][42][43][44][45][46][47][48][49]. For the purpose of having an informative cell culture model system we intended to derive primary and immortalized cell cultures from various organs of M. condylurus bats.…”

Marburg and Ebola virus infections elicit a complex, muted inflammatory state in bats

“…Viruses. Recombinant wild-type EBOV, strain Mayinga, was recovered from the full-length clone and support plasmids in HEK 293T cells and passaged twice in Vero E6 cells for amplification 29 . Recombinant wild-type MARV, strain Uganda, was recovered similarly in BHK-21 cells 89 and passaged twice in Vero E6 cells for amplification.…”

“…Some studies claim both EBOV and MARV replicate to similar levels in ERB and human derived cell lines 26 , with a robust innate immune response mounted by ERB and to a lesser degree, human cells, while others claim MARV inhibited the antiviral program in ERB cells, like in primate cells, and did not induce almost any IFN gene 27 , or little anti-viral gene induction 28 . An experiment with the pig (PK15A) and bat (EhKiT) cells suggested they responded to EBOV through the upregulation of immune, inflammatory, and coagulation pathway, in contrast to a limited response in the human (HEK293T) cells 29 . To comprehensively understand the pathways involved in the bat filoviral response, we infected bats, rather than their isolated cells, and analyzed tissue-specific RNA expression through mRNA-seq in the organs of the infected animals.…”