IFNAR2-dependent gene expression profile induced by IFN-α in Pteropus alecto bat cells and impact of IFNAR2 knockout on virus infection
August
IFNAR2-dependent gene expression profile induced by IFN-α in Pteropus alecto bat cells and impact of IFNAR2 knockout on virus infection
Qian Zhang 0 1
Lei-Ping Zeng 0 1
Peng Zhou 0 1
Aaron T. Irving 1
Shang Li 1
Zheng-Li Shi 0 1
Lin-Fa Wang 1
0 Key Laboratory of Special Pathogens, Wuhan Institute of Virology, Chinese Academy of Sciences , Wuhan, China, 2 Programme in Emerging Infectious Diseases , Duke±National University of Singapore Medical School, Singapore, Singapore, 3 Programme in Cancer and Stem Cell Biology, Duke±National University of Singapore Medical School, Singapore , Singapore
1 Editor: Eliane F Meurs, Institut Pasteur , FRANCE
Bats are important reservoirs of many viruses, which are capable of infecting the host without inducing obvious clinical diseases. Interferon and the downstream interferon regulated genes (IRGs) are known to act as the first line of defense against viral infections. Little is known about the transcriptional profile of genes being induced by interferon in bats and their role in controlling virus infection. In this study, we constructed IFNAR2 knockout bat cell lines using CRISPR technology and further characterized gene expression profiles induced by the most abundant IFN-α (IFN-α3). Firstly, we demonstrated that the CRISPR/Cas9 system is applicable for bat cells as this represents the first CRIPSR knockout cell line for bats. Our results showed the pleiotropic effect of IFN-α3 on the bat kidney cell line, PaKiT03. As expected, we confirmed that IFNAR2 is indispensable for IFN-a signaling pathway and plays an important role in antiviral immunity. Unexpectedly, we also identified novel IFNAR2-dependent IRGs which are enriched in pathways related to cancer. To our knowledge, this seems to be bat-specific as no such observation has been reported for other mammalian species. This study expands our knowledge about bat immunology and the cell line established can provide a powerful tool for future study into virus-bat interaction and cancer biology.
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Data Availability Statement: All relevant data are
within the paper and its Supporting Information
files. We have deposited the deep-sequencing
dataset in NIH Short Read Archive with the
accession number is SRR5722761.
Funding: This study was supported in part by
National Natural Science Foundation of China
(31621061) (to Z-LS) and the National Research
Foundation-Competitive Research Program
(NRFCRP) grant (NRF2012NRF-CRP001±056) from the
National Research Foundation in Singapore (to
LFW). The funders had no role in study design, data
Introduction
Most emerging and re-emerging infectious diseases are zoonotic. Bats are generally recognized
as one of the most important reservoirs of zoonotic agents and they can carry many viruses
without showing obvious clinical symptoms [1±3]. The mechanisms of how bats successfully
co-exist with various viruses are intriguing and largely remain unknown. In recent years, some
interesting findings seem to point to some bat-specific evolutionary events which resulted in
collection and analysis, decision to publish, or
preparation of the manuscript.
several bat-specific functional characteristics not shared by other mammals. Comparative
genomic analysis of Pteropus alecto and Myotis davidii shows an unexpected amount of
positively selected genes involved in the DNA damage checkpoint and NF-kB pathways [
4
]. Ahn
et al. found the functions of PYHIN gene family in bats seemed to be uniquely lost by genomic
analysis of ten bat species, which may limit excessive inflammatory activation from DNA
damage [
5
]. Although most interferon related genes are highly conserved between bats and other
mammalian species, bats exhibit some qualitative and quantitative differences in the innate
immune system [
6, 7
]. P.alecto only have three functional IFN-α genes, contrary to other
mammals which have 7±18. Moreover, P.alecto IFN-α genes seem to be constitutively
expressed in unstimulated tissues and cells, possibly acting as a 24/7 front line defense against
infection and potentially other diseases[8].
IFN-α initiates signaling through a heterodimeric transmembrane receptor termed the
IFN-α receptor (IFNAR), which is composed of IFNAR1 and IFNAR2 subunits. All IFNs can
rapidly bind to high-affinity IFNAR2 subunit, and then recruit the low-affinity IFNAR1 chain
to form an active ternary complex in human [
9
]. This process also brings into proximity the
intracellular signaling adaptors Tyk2 (with IFNAR1) and JAK1 (with IFNAR2). JAK
phosphorylate each other and further activate signal transducer and activator of transcription 1
(STAT1) and STAT2 molecules, leading to their dissociation, dimerization and finally binding
of these molecules to IRF9 to form the ISG factor 3 (ISGF3) complex. This complex then
translocates to nucleus, binds to IFN-stimulated response elements (ISRE) in interferon regulated
gene (IRG) promoters and drives the activation of IRGs transcription [
10
]. IRGs encode dir (...truncated)