Leveraging Novel Integrated Single-Cell Analyses to Define HIV-1 Latency Reversal

Zhao, Suhui; Tsibris, Athe

doi:10.3390/v13071197

Cited by 3 publications

(3 citation statements)

References 133 publications

(161 reference statements)

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“…The application of scRNA-seq in HIV research has revealed the heterogeneity in latent and reactivated HIV-1-infected cells (60)(61)(62). It has been demonstrated that latently infected CD4+ T-cells (untreated) display two cell clusters (Cluster 1 and Cluster 2) (63).…”

Section: Identification Of the Inducible Latent Cell And Potential La...mentioning

confidence: 99%

Single-Cell Sequencing Facilitates Elucidation of HIV Immunopathogenesis: A Review of Current Literature

2022

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Knowledge gaps remain in the understanding of HIV disease establishment and progression. Scientists continue to strive in their endeavor to elucidate the precise underlying immunopathogenic mechanisms of HIV-related disease, in order to identify possible preventive and therapeutic targets. A useful tool in the quest to reveal some of the enigmas related to HIV infection and disease is the single-cell sequencing (scRNA-seq) technique. With its proven capacity to elucidate critical processes in cell formation and differentiation, to decipher critical hematopoietic pathways, and to understand the regulatory gene networks that predict immune function, scRNA-seq is further considered to be a potentially useful tool to explore HIV immunopathogenesis. In this article, we provide an overview of single-cell sequencing platforms, before delving into research findings gleaned from the use of single cell sequencing in HIV research, as published in recent literature. Finally, we describe two important avenues of research that we believe should be further investigated using the single-cell sequencing technique.

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Section: Identification Of the Inducible Latent Cell And Potential La...mentioning

confidence: 99%

Single-Cell Sequencing Facilitates Elucidation of HIV Immunopathogenesis: A Review of Current Literature

2022

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“…18 Single-cell RNA-sequencing (RNA-seq), DNA-seq, or assay for transposase-accessible chromatin using sequencing (scATAC-seq) combined with new computational methods and single-cell analysis have also improved the understanding of the mechanisms of HIV latency reversal. 16,19,20 A key gap in all of the above technologies is the inability to perform time lapse imaging of single reactivating cells within a population followed by single-cell gene expression with linkage of the microscopy-measured reactivation kinetics to that cell's gene expression profile.…”

Section: Introductionmentioning

confidence: 99%

Automated microarray for single-cell sorting and collection of lymphocytes following HIV reactivation

Cortés‐Llanos

Jain

Volkheimer

et al. 2023

Preprint

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A promising strategy to cure HIV infected individuals is to use latency reversing agents (LRAs) to reactivate latent viruses, followed by host clearance of infected reservoir cells. However, reactivation of latent proviruses within infected cells is heterogeneous and often incomplete. This fact limits strategies to cure HIV which may require complete elimination of viable virus from all cellular reservoirs. For this reason, understanding the mechanism(s) of reactivation of HIV within cellular reservoirs is critical to achieve therapeutic success. Methodologies enabling temporal tracking of single cells as they reactivate followed by sorting and molecular analysis of those cells are urgently needed. To this end, microraft arrays were adapted to image T-lymphocytes expressing mCherry under the control of the HIV long terminal repeat (LTR) promoter, in response to the application of various LRAs (prostratin, iBET151, and SAHA). In response to prostratin, iBET151, and SAHA, 30.5 %, 11.2 %, and 12.1 % percentage of cells respectively, reactivated similar to that observed in other experimental systems. The arrays enabled large numbers of single cells (>25,000) to be imaged over time. mCherry fluorescence quantification identified cell subpopulations with differing reactivation kinetics. Significant heterogeneity was observed at the single cell level between different LRAs in terms of time to reactivation, rate of mCherry fluorescence increase upon reactivation, and peak fluorescence attained. In response to prostratin, subpopulations of T lymphocytes with slow and fast reactivation kinetics were identified. Single T-lymphocytes that were either fast or slow reactivators were sorted, and single-cell RNA-sequencing was performed. Different genes associated with inflammation, immune activation, and cellular and viral transcription factors were found. These results advance our conceptual understanding of HIV reactivation dynamics at the single-cell level toward a cure for HIV.

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“…Of these methods, time‐lapse imaging of single cells engineered with fluorescent reporters has been particularly valuable in understanding heterogeneity in reactivation kinetics 18 . Single‐cell RNA‐sequencing (scRNA‐seq), DNA‐seq, or assay for transposase‐accessible chromatin using sequencing (scATAC‐seq) combined with new computational methods and single‐cell analysis have also improved the understanding of the mechanisms of HIV latency reversal 16,19,20 . A key gap in all of the above technologies is the inability to perform time‐lapse imaging of single reactivating cells within a population followed by single‐cell gene expression with linkage of the microscopy‐measured reactivation kinetics to that cell's gene expression profile.…”

Section: Introductionmentioning

confidence: 99%

Automated microarray platform for single‐cell sorting and collection of lymphocytes following HIV reactivation

Cortés‐Llanos

Jain

Volkheimer

et al. 2023

Bioengineering & Transla Med

View full text Add to dashboard Cite

A promising strategy to cure HIV‐infected individuals is to use latency reversing agents (LRAs) to reactivate latent viruses, followed by host clearance of infected reservoir cells. However, reactivation of latent proviruses within infected cells is heterogeneous and often incomplete. This fact limits strategies to cure HIV which may require complete elimination of viable virus from all cellular reservoirs. For this reason, understanding the mechanism(s) of reactivation of HIV within cellular reservoirs is critical to achieve therapeutic success. Methodologies enabling temporal tracking of single cells as they reactivate followed by sorting and molecular analysis of those cells are urgently needed. To this end, microraft arrays were adapted to image T‐lymphocytes expressing mCherry under the control of the HIV long terminal repeat (LTR) promoter, in response to the application of LRAs (prostratin, iBET151, and SAHA). In response to prostratin, iBET151, and SAHA, 30.5%, 11.2%, and 12.1% percentage of cells, respectively. The arrays enabled large numbers of single cells (>25,000) to be imaged over time. mCherry fluorescence quantification identified cell subpopulations with differing reactivation kinetics. Significant heterogeneity was observed at the single‐cell level between different LRAs in terms of time to reactivation, rate of mCherry fluorescence increase upon reactivation, and peak fluorescence attained. In response to prostratin, subpopulations of T lymphocytes with slow and fast reactivation kinetics were identified. Single T‐lymphocytes that were either fast or slow reactivators were sorted, and single‐cell RNA‐sequencing was performed. Different genes associated with inflammation, immune activation, and cellular and viral transcription factors were found.

show abstract

Leveraging Novel Integrated Single-Cell Analyses to Define HIV-1 Latency Reversal

Cited by 3 publications

References 133 publications

Single-Cell Sequencing Facilitates Elucidation of HIV Immunopathogenesis: A Review of Current Literature

Single-Cell Sequencing Facilitates Elucidation of HIV Immunopathogenesis: A Review of Current Literature

Automated microarray for single-cell sorting and collection of lymphocytes following HIV reactivation

Automated microarray platform for single‐cell sorting and collection of lymphocytes following HIV reactivation

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