A Dual-reporter system for real-time monitoring and high-throughput CRISPR/Cas9 library screening of the hepatitis C virus.
The hepatitis C virus (HCV) is one of the leading causes of chronic hepatitis, liver cirrhosis and hepatocellular carcinomas and infects approximately 170 million people worldwide. Although several reporter systems have been developed, many shortcomings limit their use in the assessment of HCV infections. Here, we report a real-time live-cell reporter, termed the NIrD (NS3-4A Inducible rtTA-mediated Dual-reporter) system, which provides an on-off switch specifically in response to an HCV infection. Using the NIrD system and a focused CRISPR/Cas9 library, we identified CLDN1, OCLN and CD81 as essential genes for both the cell-free entry and the cell-to-cell transmission of HCV. The combination of this ultra-sensitive reporter system and the CRISPR knockout screening provides a powerful and high-throughput strategy for the identification of critical host components for HCV infections.
Biodynamic Optical Imaging Center (BIOPIC), Peking-Tsinghua Center for Life Sciences, State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences, Peking University, Beijing 100871, China.
Qingpeng
Q
Ren
Biodynamic Optical Imaging Center (BIOPIC), Peking-Tsinghua Center for Life Sciences, State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences, Peking University, Beijing 100871, China.
Chan
C
Li
Biodynamic Optical Imaging Center (BIOPIC), Peking-Tsinghua Center for Life Sciences, State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences, Peking University, Beijing 100871, China.
Pengfei
P
Yuan
Biodynamic Optical Imaging Center (BIOPIC), Peking-Tsinghua Center for Life Sciences, State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences, Peking University, Beijing 100871, China.
Changzu
C
Cai
Comprehensive AIDS Research Center and Research Center for Public Health, School of Medicine, Tsinghua University, Beijing 100084, China.
Linqi
L
Zhang
1] Department of Microbiology, University of Alabama at Birmingham School of Medicine, Birmingham, Alabama 35294, USA [2] Department of Microbiology, Peking University College of Basic Medical Sciences, Beijing 100083, China.
Guangxiang George
GG
Luo
Biodynamic Optical Imaging Center (BIOPIC), Peking-Tsinghua Center for Life Sciences, State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences, Peking University, Beijing 100871, China.
Wensheng
W
Wei
D064112:Clustered Regularly Interspaced Short Palindromic Repeats / Q000235:genetics*; D015723:Gene Library; D017930:Genes, Reporter / Q000235:genetics*; D006367:HeLa Cells; D016174:Hepacivirus; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans; D013050:Spectrometry, Fluorescence / Q000379:methods; D014764:Viral Proteins / Q000235:genetics*; D053586:Virus Internalization
D014764:Viral Proteins