NIH grant: In Situ Structural Biology of HIV-1 Transcription and Latency in CD4⁺ T Cells (1R21AI200512-01)

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NIH grant 1R21AI200512-01: In Situ Structural Biology of HIV-1 Transcription and Latency in CD4⁺ T Cells
Institute: {'code': 'AI', 'abbreviation': 'NIAID', 'name': 'National Institute of Allergy and Infectious Diseases', 'admin_org_id': '12500003', 'admin_funding_url': ''}. PI: Peijun  Zhang. Award: $170980.
Start: 2026-06-23T00:00:00.
Project Summary
Human immunodeficiency virus type 1 (HIV-1) persists in long-lived CD4⁺ T cells despite
effective antiretroviral therapy, where integrated proviruses remain silent or intermittently
active. This latent reservoir is the major barrier to cure. Whether a provirus is latent or
productive depends on its nuclear environment, including chromatin organization, proximity
to nuclear bodies, positioning of nucleosomes, and access to transcriptional machinery.
Current knowledge from the past 30 years of study is based largely on bulk assays that
average across cells and require artificial enrichment of “latent” populations, masking the
true heterogeneity of proviral states.
Microscopy-based structural biology offers a unique solution. Correlative light and electron
microscopy (CLEM) and cryo-electron tomography (cryo-ET) enable direct visualization of
individual proviruses in intact nuclei, allowing transcriptionally active and silent proviruses to
be examined side by side. This approach provides a population-based nanoscale view of
latency and active transcription, revealing diverse structural states.
The goal of this project is to define the structural basis of HIV persistence. We will apply
cryo-CLEM, cryo-focused ion beam (cryo-FIB) milling, and cryo-ET to visualize the nuclear
environments of HIV proviruses in both CEM cell lines and primary CD4⁺ T cells. We
hypothesize that nanoscale chromatin and nuclear body architecture determines proviral
competence and that distinct latency states are structurally distinguishable and remodelable.
Aim 1 will establish dual-reporter strategies in CEM cells to label proviral DNA and nascent
RNA, enabling identification of active and silent proviruses and mapping of their nuclear
niches by live-cell imaging and correlative cryo-ET. Aim 2 will extend these methods to
physiologically relevant primary CD4⁺ T cells, classifying distinct nuclear structural features
associated with latency and transcription.
This project will deliver the first in situ structural characterization of HIV proviruses in primary
CD4⁺ T cells. The results will reveal nanoscale determinants of reservoir stability, clarify how
nuclear architecture regulates proviral activity, and inform rational cure strategies. More
broadly, the work will establish imaging approaches generally applicable to transcriptional
regulation in both viral and host systems.

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