title: Specific Reactivation of Latent HIV-1 by dCas9-SunTag-VP64-mediated Guide RNA Targeting the HIV-1 Promoter.
abstract: HIV-1 escapes antiretroviral agents by integrating into the host DNA and forming a latent transcriptionally silent HIV-1 provirus. Transcriptional activation is prerequisite for reactivation and the eradication of latent HIV-1 proviruses. dCas9-SunTag-VP64 transcriptional system has been reported that it can robustly activate the expression of an endogenous gene using a single guide RNA (sgRNA). Here, we systematically investigated the potential of dCas9-SunTag-VP64 with the designed sgRNAs for reactivating latent HIV-1. We found dCas9-SunTag-VP64 with sgRNA 4 or sgRNA 5 targeted from -164 to -146 or -124 to -106 bp upstream of the transcription start sites of HIV-1 could induce high expression of luciferase reporter gene after screening of sgRNAs targeting different regions of the HIV-1 promoter. Further, we confirmed that dCas9-SunTag-VP64 with sgRNA 4 or sgRNA 5 can effectively reactivate latent HIV-1 transcription in several latently infected human T-cell lines. Moreover, we confirmed that the reactivation of latent HIV-1 by dCas9-SunTag-VP64 with the designed sgRNA occurred through specific binding to the HIV-1 LTR promoter without genotoxicity and global T-cell activation. Taken together, our data demonstrated dCas9-SunTag-VP64 system can effectively and specifically reactivate latent HIV-1 transcription, suggesting that this strategy could offer a novel approach to anti-HIV-1 latency.
authors.affiliation: State Key Laboratory of Genetic Engineering, and Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, School of Life Sciences, Fudan University, Shanghai, China.
authors.forename: Haiyan
authors.initials: H
authors.lastname: Ji
authors.affiliation: State Key Laboratory of Genetic Engineering, and Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, School of Life Sciences, Fudan University, Shanghai, China.
authors.forename: Zhengtao
authors.initials: Z
authors.lastname: Jiang
authors.affiliation: State Key Laboratory of Genetic Engineering, and Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, School of Life Sciences, Fudan University, Shanghai, China.
authors.forename: Panpan
authors.initials: P
authors.lastname: Lu
authors.affiliation: Key Laboratory of Molecular Virology and Immunology, Institute Pasteur of Shanghai, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China.
authors.forename: Li
authors.initials: L
authors.lastname: Ma
authors.affiliation: Key Laboratory of Molecular Virology and Immunology, Institute Pasteur of Shanghai, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China.
authors.forename: Chuan
authors.initials: C
authors.lastname: Li
authors.affiliation: State Key Laboratory of Genetic Engineering, and Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, School of Life Sciences, Fudan University, Shanghai, China.
authors.forename: Hanyu
authors.initials: H
authors.lastname: Pan
authors.affiliation: State Key Laboratory of Genetic Engineering, and Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, School of Life Sciences, Fudan University, Shanghai, China.
authors.forename: Zheng
authors.initials: Z
authors.lastname: Fu
authors.affiliation: State Key Laboratory of Genetic Engineering, and Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, School of Life Sciences, Fudan University, Shanghai, China.
authors.forename: Xiying
authors.initials: X
authors.lastname: Qu
authors.affiliation: State Key Laboratory of Genetic Engineering, and Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, School of Life Sciences, Fudan University, Shanghai, China.
authors.forename: Pengfei
authors.initials: P
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Genetic Engineering, and Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, School of Life Sciences, Fudan University, Shanghai, China.
authors.forename: Junxiao
authors.initials: J
authors.lastname: Deng
authors.affiliation: State Key Laboratory of Genetic Engineering, and Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, School of Life Sciences, Fudan University, Shanghai, China.
authors.forename: Xinyi
authors.initials: X
authors.lastname: Yang
authors.affiliation: Key Laboratory of Molecular Virology and Immunology, Institute Pasteur of Shanghai, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China.
authors.forename: Jianhua
authors.initials: J
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Genetic Engineering, and Key Laboratory of Medical Molecular Virology of Ministry of Education/Health, School of Life Sciences, Fudan University, Shanghai, China.
authors.forename: Huanzhang
authors.initials: H
authors.lastname: Zhu
mesh_terms: D001483:Base Sequence; D001665:Binding Sites; D064113:CRISPR-Cas Systems; D002460:Cell Line; D015870:Gene Expression; D015967:Gene Expression Regulation, Viral; D023061:Gene Order; D018390:Gene Targeting; D017930:Genes, Reporter; D005822:Genetic Vectors / Q000235:genetics; D016325:HIV Long Terminal Repeat; D015497:HIV-1 / Q000502:physiology*; D006801:Humans; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011993:Recombinant Fusion Proteins; D015533:Transcriptional Activation; D014775:Virus Activation / Q000235:genetics*
chemical_list: D011993:Recombinant Fusion Proteins