Designed Transcription Activator-Like Effector Proteins Efficiently Induced the Expression of Latent HIV-1 in Latently Infected Cells

Wang, Xiaohui; Wang, Pengfei; Fu, Zheng; Ji, Haiyan; Qu, Xin; Zeng, Hulie; Zhu, Xiaoli; Deng, Jixin; Lu, Panpan; Zha, Shijun; Song, Zhishuo; Zhu, Huanzhang

doi:10.1089/aid.2014.0121

Cited by 15 publications

(18 citation statements)

References 43 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The presence of residual latent but replication-competent HIV-1 reservoirs is a major hurdle impeding viral eradication. Recent work has indicated that HIV expression can be induced using zincfinger 83 and TAL effector-based 84 transcription factors engineered to bind the LTR promoter. Here, we show that CRISPR/Cas9based transcriptional effectors-which can be retargeted to nearly any DNA sequence without protein engineering-can reactivate viral gene expression in cell line models of HIV-1 latency.…”

Section: Discussionmentioning

confidence: 99%

CRISPR-mediated Activation of Latent HIV-1 Expression

2016

View full text Add to dashboard Cite

Complete eradication of HIV-1 infection is impeded by the existence of cells that harbor chromosomally integrated but transcriptionally inactive provirus. These cells can persist for years without producing viral progeny, rendering them refractory to immune surveillance and antiretroviral therapy and providing a permanent reservoir for the stochastic reactivation and reseeding of HIV-1. Strategies for purging this latent reservoir are thus needed to eradicate infection. Here, we show that engineered transcriptional activation systems based on CRISPR/Cas9 can be harnessed to activate viral gene expression in cell line models of HIV-1 latency. We further demonstrate that complementing Cas9 activators with latency-reversing compounds can enhance latent HIV-1 transcription and that epigenome modulation using CRISPR-based acetyltransferases can also promote viral gene activation. Collectively, these results demonstrate that CRISPR systems are potentially effective tools for inducing latent HIV-1 expression and that their use, in combination with antiretroviral therapy, could lead to improved therapies for HIV-1 infection.

show abstract

Section: Discussionmentioning

confidence: 99%

CRISPR-mediated Activation of Latent HIV-1 Expression

2016

View full text Add to dashboard Cite

show abstract

“…However, it cannot eliminate the integrated HIV-1 genome, and CCR5 is not the sole receptor for HIV-1 infection and has many other cellular functions as well[20]. The highly conserved HIV-1 LTR has been targeted by ZFN and TALEN for the eradication of the integrated HIV-1 genome[24-26]. The major drawbacks for ZFNs and TALENs are the tedious engineering of custom DNA-binding fusion proteins for each target site, which limit their universal application and clinical safety[27-30].…”

Section: Introductionmentioning

confidence: 99%

Functional screening of guide RNAs targeting the regulatory and structural HIV-1 viral genome for a cure of AIDS

Yin

Zhang

et al. 2016

Aids

View full text Add to dashboard Cite

Objective There is an urgent need for the development of HIV-1 genome eradication strategies that lead to a permanent cure for HIV-1/AIDS. We previously reported that 4 guide RNAs (gRNAs) targeting HIV-1 long terminal repeats (LTR) effectively eradicated the entire HIV-1 genome. In this study, we sought to identify the best gRNAs targeting HIV-1 LTR and viral structural region and to optimize gRNA pairing that can efficiently eradicate the HIV-1 genome. Design Highly specific gRNAs were designed using bioinformatics tools and their capacity of guiding Cas9 to cleave HIV-1 proviral DNA was evaluated using high throughput HIV-1 luciferase reporter assay and rapid Direct-PCR genotyping. Methods The target seed sequences for each gRNA were cloned into lentiviral vectors. HEK293T cells were cotransfected with a pEcoHIV-NL4-3-firefly-luciferase reporter vector (1:20) over lentiviral vectors carrying Cas9 and single gRNA or various combinations of gRNAs. The EcoHIV DNA cleaving efficiency was evaluated by Direct-PCR genotyping and the EcoHIV transcription/replication activity was examined by a luciferase reporter assay. Results Most of the designed gRNAs are effective to eliminate the predicted HIV-1 genome sequence between the selected two target sites. This is evidenced by the presence of PCR genotypic deletion/insertion and the decrease of luciferase reporter activity. In particular, a combination of viral structural gRNAs with LTR gRNAs provided a higher efficiency of genome eradication and an easier approach for PCR genotyping. Conclusion Our screening strategy can specifically and effectively identify gRNAs targeting HIV-1 LTR and structural region to excise proviral HIV-1 from the host genome.

show abstract

“…Like ZFPs, TALENs have been used to knock out the CCR5 gene in cell culture [203][204][205][206], permitting efficient gene editing and protection of cells from HIV infection [207] with minimal off-target activity and low cytotoxicity. TALE-activators, corresponding to TALEs fused to potent transcription activators, have been developed and shown to effectively reactivate latent HIV-1 in cell line models [208][209][210] and in PBMCs from ART-suppressed individuals [210]. TALE technology combines several features that are quite favorable for gene editing applications.…”

Section: Talesmentioning

confidence: 99%

Reduce and Control: A Combinatorial Strategy for Achieving Sustained HIV Remissions in the Absence of Antiretroviral Therapy

Schwarzer

Gramatica

Greene

2020

Viruses

View full text Add to dashboard Cite

Human immunodeficiency virus (HIV-1) indefinitely persists, despite effective antiretroviral therapy (ART), within a small pool of latently infected cells. These cells often display markers of immunologic memory and harbor both replication-competent and -incompetent proviruses at approximately a 1:100 ratio. Although complete HIV eradication is a highly desirable goal, this likely represents a bridge too far for our current and foreseeable technologies. A more tractable goal involves engineering a sustained viral remission in the absence of ART--a "functional cure." In this setting, HIV remains detectable during remission, but the size of the reservoir is small and the residual virus is effectively controlled by an engineered immune response or other intervention. Biological precedence for such an approach is found in the post-treatment controllers (PTCs), a rare group of HIV-infected individuals who, following ART withdrawal, do not experience viral rebound. PTCs are characterized by a small reservoir, greatly reduced inflammation, and the presence of a poorly understood immune response that limits viral rebound. Our goal is to devise a safe and effective means for replicating durable post-treatment control on a global scale. This requires devising methods to reduce the size of the reservoir and to control replication of this residual virus. In the following sections, we will review many of the approaches and tools that likely will be important for implementing such a "reduce and control" strategy and for achieving a PTC-like sustained HIV remission in the absence of ART.

show abstract

Designed Transcription Activator-Like Effector Proteins Efficiently Induced the Expression of Latent HIV-1 in Latently Infected Cells

Cited by 15 publications

References 43 publications

CRISPR-mediated Activation of Latent HIV-1 Expression

CRISPR-mediated Activation of Latent HIV-1 Expression

Functional screening of guide RNAs targeting the regulatory and structural HIV-1 viral genome for a cure of AIDS

Reduce and Control: A Combinatorial Strategy for Achieving Sustained HIV Remissions in the Absence of Antiretroviral Therapy

Contact Info

Product

Resources

About