학술논문
CRISPR-Cas9 screen of E3 ubiquitin ligases identifies TRAF2 and UHRF1 as regulators of HIV latency in primary human T cells.
Document Type
Academic Journal
Author
Rathore U; Gladstone Institutes, San Francisco, California, USA.; Department of Microbiology and Immunology, University of California, San Francisco, California, USA.; Innovative Genomics Institute, University of California, Berkeley, California, USA.; Haas P; Gladstone Institutes, San Francisco, California, USA.; Quantitative Biosciences Institute (QBI), University of California, San Francisco, California, USA.; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, California, USA.; Easwar Kumar V; Gladstone Institutes, San Francisco, California, USA.; Department of Microbiology and Immunology, University of California, San Francisco, California, USA.; Innovative Genomics Institute, University of California, Berkeley, California, USA.; Hiatt J; Gladstone Institutes, San Francisco, California, USA.; Department of Microbiology and Immunology, University of California, San Francisco, California, USA.; Innovative Genomics Institute, University of California, Berkeley, California, USA.; Medical Scientist Training Program, University of California, San Francisco, California, USA.; Biomedical Sciences Graduate Program, University of California, San Francisco, California, USA.; Haas KM; Gladstone Institutes, San Francisco, California, USA.; Quantitative Biosciences Institute (QBI), University of California, San Francisco, California, USA.; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, California, USA.; Bouhaddou M; Gladstone Institutes, San Francisco, California, USA.; Quantitative Biosciences Institute (QBI), University of California, San Francisco, California, USA.; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, California, USA.; Swaney DL; Gladstone Institutes, San Francisco, California, USA.; Quantitative Biosciences Institute (QBI), University of California, San Francisco, California, USA.; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, California, USA.; Stevenson E; Gladstone Institutes, San Francisco, California, USA.; Quantitative Biosciences Institute (QBI), University of California, San Francisco, California, USA.; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, California, USA.; Zuliani-Alvarez L; Gladstone Institutes, San Francisco, California, USA.; Quantitative Biosciences Institute (QBI), University of California, San Francisco, California, USA.; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, California, USA.; McGregor MJ; Gladstone Institutes, San Francisco, California, USA.; Quantitative Biosciences Institute (QBI), University of California, San Francisco, California, USA.; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, California, USA.; Turner-Groth A; Gladstone Institutes, San Francisco, California, USA.; Ochieng' Olwal C; West African Centre for Cell Biology of Infectious Pathogens (WACCBIP), College of Basic and Applied Sciences, University of Ghana, Accra, Ghana.; Department of Biochemistry, Cell & Molecular Biology, College of Basic & Applied Sciences, University of Ghana, Accra, Ghana.; Bediako Y; West African Centre for Cell Biology of Infectious Pathogens (WACCBIP), College of Basic and Applied Sciences, University of Ghana, Accra, Ghana.; Department of Biochemistry, Cell & Molecular Biology, College of Basic & Applied Sciences, University of Ghana, Accra, Ghana.; Braberg H; Gladstone Institutes, San Francisco, California, USA.; Quantitative Biosciences Institute (QBI), University of California, San Francisco, California, USA.; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, California, USA.; Soucheray M; Gladstone Institutes, San Francisco, California, USA.; Quantitative Biosciences Institute (QBI), University of California, San Francisco, California, USA.; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, California, USA.; Ott M; Gladstone Institutes, San Francisco, California, USA.; Eckhardt M; Gladstone Institutes, San Francisco, California, USA.; Quantitative Biosciences Institute (QBI), University of California, San Francisco, California, USA.; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, California, USA.; Hultquist JF; Division of Infectious Diseases, Northwestern University Feinberg School of Medicine, Chicago, Illinois, USA.; Center for Pathogen Genomics and Microbial Evolution, Institute for Global Health, Northwestern University Feinberg School of Medicine, Chicago, Illinois, USA.; Marson A; Gladstone Institutes, San Francisco, California, USA.; Department of Microbiology and Immunology, University of California, San Francisco, California, USA.; Innovative Genomics Institute, University of California, Berkeley, California, USA.; Department of Medicine, University of California, San Francisco, California, USA.; Diabetes Center, University of California, San Francisco, California, USA.; UCSF Helen Diller Family Comprehensive Cancer Center, University of California, San Francisco, California, USA.; Parker Institute for Cancer Immunotherapy, University of California, San Francisco, California, USA.; Institute for Human Genetics, University of California, San Francisco, California, USA.; Kaake RM; Gladstone Institutes, San Francisco, California, USA.; Quantitative Biosciences Institute (QBI), University of California, San Francisco, California, USA.; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, California, USA.; Krogan NJ; Gladstone Institutes, San Francisco, California, USA.; Quantitative Biosciences Institute (QBI), University of California, San Francisco, California, USA.; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, California, USA.
Source
Publisher: American Society for Microbiology Country of Publication: United States NLM ID: 101519231 Publication Model: Print-Electronic Cited Medium: Internet ISSN: 2150-7511 (Electronic) NLM ISO Abbreviation: mBio Subsets: MEDLINE
Subject
Language
English
Abstract
During HIV infection of CD4+ T cells, ubiquitin pathways are essential to viral replication and host innate immune response; however, the role of specific E3 ubiquitin ligases is not well understood. Proteomics analyses identified 116 single-subunit E3 ubiquitin ligases expressed in activated primary human CD4+ T cells. Using a CRISPR-based arrayed spreading infectivity assay, we systematically knocked out 116 E3s from activated primary CD4+ T cells and infected them with NL4-3 GFP reporter HIV-1. We found 10 E3s significantly positively or negatively affected HIV infection in activated primary CD4+ T cells, including UHRF1 (pro-viral) and TRAF2 (anti-viral). Furthermore, deletion of either TRAF2 or UHRF1 in three JLat models of latency spontaneously increased HIV transcription. To verify this effect, we developed a CRISPR-compatible resting primary human CD4+ T cell model of latency. Using this system, we found that deletion of TRAF2 or UHRF1 initiated latency reactivation and increased virus production from primary human resting CD4+ T cells, suggesting these two E3s represent promising targets for future HIV latency reversal strategies.
Importance: HIV, the virus that causes AIDS, heavily relies on the machinery of human cells to infect and replicate. Our study focuses on the host cell's ubiquitination system which is crucial for numerous cellular processes. Many pathogens, including HIV, exploit this system to enhance their own replication and survival. E3 proteins are part of the ubiquitination pathway that are useful drug targets for host-directed therapies. We interrogated the 116 E3s found in human immune cells known as CD4+ T cells, since these are the target cells infected by HIV. Using CRISPR, a gene-editing tool, we individually removed each of these enzymes and observed the impact on HIV infection in human CD4+ T cells isolated from healthy donors. We discovered that 10 of the E3 enzymes had a significant effect on HIV infection. Two of them, TRAF2 and UHRF1, modulated HIV activity within the cells and triggered an increased release of HIV from previously dormant or "latent" cells in a new primary T cell assay. This finding could guide strategies to perturb hidden HIV reservoirs, a major hurdle to curing HIV. Our study offers insights into HIV-host interactions, identifies new factors that influence HIV infection in immune cells, and introduces a novel methodology for studying HIV infection and latency in human immune cells.
Importance: HIV, the virus that causes AIDS, heavily relies on the machinery of human cells to infect and replicate. Our study focuses on the host cell's ubiquitination system which is crucial for numerous cellular processes. Many pathogens, including HIV, exploit this system to enhance their own replication and survival. E3 proteins are part of the ubiquitination pathway that are useful drug targets for host-directed therapies. We interrogated the 116 E3s found in human immune cells known as CD4+ T cells, since these are the target cells infected by HIV. Using CRISPR, a gene-editing tool, we individually removed each of these enzymes and observed the impact on HIV infection in human CD4+ T cells isolated from healthy donors. We discovered that 10 of the E3 enzymes had a significant effect on HIV infection. Two of them, TRAF2 and UHRF1, modulated HIV activity within the cells and triggered an increased release of HIV from previously dormant or "latent" cells in a new primary T cell assay. This finding could guide strategies to perturb hidden HIV reservoirs, a major hurdle to curing HIV. Our study offers insights into HIV-host interactions, identifies new factors that influence HIV infection in immune cells, and introduces a novel methodology for studying HIV infection and latency in human immune cells.