ABSTRACT
Bats are reservoirs for numerous viruses that cause serious diseases in other animals and humans. Several mechanisms are proposed to contribute to the tolerance of bats to these pathogens. This study investigates the response of bat cells to double-stranded RNA generated by SARS-CoV-2 replication. Here, we found the involvement of Dicer in the processing of viral genomic RNAs during SARS-CoV-2 infection. Examining RNA sequencing of infected cells, small-interfering RNA (siRNA)-like fragments were found derived from viral RNAs. Depletion of Dicer showed a reduction in these RNAs and an increase in viral loads suggesting unlike other mammals, bats may use Dicer to limit viral replication. This prompted the exploration of key dsRNA sensors in bat cells. Our analysis showed significant upregulation of OAS1 and MX1 in response to dsRNA, while PKR levels remained low, suggesting alternative dsRNA-response mechanisms are present that eschew the common PKR-based system. These results further show how bats employ distinct strategies for antiviral defense that may contribute to tolerating viral infections. They suggest the involvement of Dicer in antiviral mechanisms in bats, a function not observed in other mammals. This highlights a mechanism for bat originating viruses to evolve features that in other animals could cause extreme antiviral responses such as is seen with SARS-CoV-2.
Competing Interest Statement
The authors have declared no competing interest.
LIST OF ABBREVIATIONS
- siRNA
- Small-interfering RNA
- OAS1
- 2’-5’-Oligoadenylate Synthetase 1
- MX1
- Myxovirus Resistance Protein 1
- dsRNA
- Double-stranded RNA
- PKR
- Protein Kinase R
- SARS-CoV-2
- Severe Acute Respiratory Syndrome Coronavirus 2
- miRNA
- microRNA
- SARS-CoV
- Severe Acute Respiratory Syndrome Coronavirus
- MERS-CoV
- Middle East Respiratory Syndrome Coronavirus
- ACE2
- Angiotensin-Converting Enzyme 2
- Spike (S)
- Spike Protein
- Envelope (E)
- Envelope Protein
- Matrix (M)
- Matrix Protein
- Nucleocapsid (N)
- Nucleocapsid Protein
- nsp
- Non-structural Protein
- HCoV-229E
- Human Coronavirus 229E
- HCoV-NL63
- Human Coronavirus NL63
- P. alecto
- Pteropus alecto
- IFN
- Type I and Type II Interferon
- NF-κB
- Nuclear Factor Kappa-light-chain-enhancer of Activated B Cells
- NLRP3
- NOD-, LRR- and Pyrin Domain-containing Protein 3
- TLRs
- Toll-like Receptors
- eIF2α
- Eukaryotic Initiation Factor 2 Alpha
- ATF4
- Activating Transcription Factor 4
- ATF3
- Activating Transcription Factor 3
- RNAi
- RNA Interference
- VSRs
- Viral Suppressors of RNA Silencing
- TBLU
- Tadarida brasiliensis (Brazilian Free-tailed Bat)
- sgRNAs
- Subgenomic RNAs
- qPCR
- Quantitative Polymerase Chain Reaction
- PCR
- Polymerase Chain Reaction
- DKD
- Double Knockdown
- MOI
- Multiplicity of Infection
- Vero E6
- Epithelial Cells Derived from African Green Monkey Kidney
- 293T
- Human Embryonic Kidney 293 Cells Expressing SV40 T-antigen
- BHK
- Baby Hamster Kidney Cells
- vsiRNAs
- Virus-derived Small-interfering RNAs
- SINV
- Sindbis Virus
- PaKi cells
- Pteropus alecto Kidney Cells
- mRNA
- messenger RNA
- GO
- Gene Ontology
- Poly(I:C)
- Polyinosinic:polycytidylic acid
- FPKM
- Fragments Per Kilobase of Transcript Per Million Mapped Reads
- HEK293
- Human Embryonic Kidney 293 Cells
- Phospho
- Phosphorylated
- Thr
- Threonine
- Ser
- Serine
- OB
- Oligonucleotide Binding
- CTD
- C-terminal Domain
- P-loop
- Phosphate-binding Loop
- COVID-19
- Coronavirus Disease 2019
- BSL-3
- Biosafety Level 3
- PFU
- Plaque Forming Units
- RT–qPCR
- Reverse Transcription Quantitative Polymerase Chain Reaction
- TBS-T
- Tris-Buffered Saline with Tween
- HRP
- Horseradish Peroxidase
- PVDF
- Polyvinylidene Fluoride
- SDS-PAGE
- Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis
- VST
- Variance Stabilizing Transformation