A high-throughput screen for genes essential for PRRSV infection using a piggyBac-based system

Jianhui Bai; Kongpan Li; Wenda Tang; Zuoxiang Liang; Xifeng Wang; Wenhai Feng; Shujun Zhang; Liming Ren; Sen Wu; Haitang Han; Yaofeng Zhao

doi:10.1016/j.virol.2019.03.001

A high-throughput screen for genes essential for PRRSV infection using a piggyBac-based system

Virology. 2019 May:531:19-30. doi: 10.1016/j.virol.2019.03.001. Epub 2019 Mar 4.

Authors

Jianhui Bai¹, Kongpan Li¹, Wenda Tang¹, Zuoxiang Liang¹, Xifeng Wang², Wenhai Feng¹, Shujun Zhang³, Liming Ren¹, Sen Wu⁴, Haitang Han⁵, Yaofeng Zhao⁶

Affiliations

¹ State Key Laboratory of Agrobiotechnology, College of Biological Sciences, National Engineering Laboratory for Animal Breeding, China Agricultural University, Beijing 100193, PR China.
² Key Laboratory of Animal Ecology and Conservation Biology, Institute of Zoology, Chinese Academy of Science, Beijing 100101, PR China.
³ Key Laboratory of Agricultural Animal Genetics, Breeding and Reproduction of Ministry of Education, Huazhong Agricultural University, Wuhan, Hubei, PR China.
⁴ State Key Laboratory of Agrobiotechnology, College of Biological Sciences, National Engineering Laboratory for Animal Breeding, China Agricultural University, Beijing 100193, PR China. Electronic address: swu@cau.edu.cn.
⁵ State Key Laboratory of Agrobiotechnology, College of Biological Sciences, National Engineering Laboratory for Animal Breeding, China Agricultural University, Beijing 100193, PR China. Electronic address: hanhaitang@cau.edu.cn.
⁶ State Key Laboratory of Agrobiotechnology, College of Biological Sciences, National Engineering Laboratory for Animal Breeding, China Agricultural University, Beijing 100193, PR China. Electronic address: yaofengzhao@cau.edu.cn.

PMID: 30852268
DOI: 10.1016/j.virol.2019.03.001

Abstract

In this study, using a dual-functional, piggyBac transposon-based system, we developed a method to systematically decipher the host genes that may be associated with porcine reproductive and respiratory syndrome virus (PRRSV) infection. A Marc145 cell library, which was randomly mutated by transfecting piggyBac plasmids, was challenged with PRRSV. The surviving cell clones were subjected to inverse PCR and high-throughput sequencing to map the integration sites of the transposon. Detailed annotation of the genes flanking the integration sites allowed us to generate a ranked list of candidate genes. Among the predicted genes with a high priority, four genes, CDK17, RNF168, BCL2L15, and TRIM33, were strongly correlated with PRRSV infection in both Marc145 cells and porcine primary alveolar macrophages. This study not only assists in identifying the genes essential for PRRSV infection but also confirms the possibility of using the piggyBac system to study other virus-host genetic interactions in a high-throughput manner.

Keywords: BCL2L15; CDK17; High-throughput screen; Porcine reproductive and respiratory syndrome virus (PRRSV); RNF168; TRIM33; piggyBac transposon.

Publication types

Evaluation Study
Research Support, Non-U.S. Gov't

MeSH terms

Animals
Cyclin-Dependent Kinases / genetics
Cyclin-Dependent Kinases / metabolism
DNA Transposable Elements
Gene Expression Profiling
High-Throughput Screening Assays / methods*
Host-Pathogen Interactions
Macrophages, Alveolar / metabolism
Macrophages, Alveolar / virology
Plasmids / genetics
Plasmids / metabolism
Porcine Reproductive and Respiratory Syndrome / genetics*
Porcine Reproductive and Respiratory Syndrome / metabolism
Porcine Reproductive and Respiratory Syndrome / virology*
Porcine respiratory and reproductive syndrome virus / genetics
Porcine respiratory and reproductive syndrome virus / physiology*
Swine
Transcription Factors / genetics
Transcription Factors / metabolism
Ubiquitin-Protein Ligases / genetics
Ubiquitin-Protein Ligases / metabolism
Virus Replication

Substances

DNA Transposable Elements
Transcription Factors
Ubiquitin-Protein Ligases
Cyclin-Dependent Kinases