In a recent paper published in the journal Molecular Cell, researchers present a valuable data resource that provides detailed insight into the cellular and genetic mechanisms that govern human immunodeficiency virus (HIV) integration and expression in the brain human
Study: HIV integration into the human brain is linked to microglia activation and 3D genome remodeling. Image credit: SquareMotion / Shutterstock.com
background
HIV enters the central nervous system (CNS) during the first two weeks after infection. Microglia, myeloid cells, and macrophages, which collectively constitute 5–10% of neurons, are primarily infected.
HIV-infected with encephalitis (HIV) is a rare complication after the discovery of combination antiretroviral therapy (cART); however, HIVE is the ideal condition to study how active HIV replication affects the brain. In some cases, during the initial stages of acute HIV infection, patients present with neurological symptoms similar to a hive-like state.
More genome- and transcriptome-level data are needed to design effective treatments for HIV-associated neurocognitive disorder (HAND), which affects 20–50% of HIV-infected people worldwide. To date, most genomic studies have focused on gene expression profiling of bulk tissues to demonstrate HIV-induced metabolic changes and neuroinflammation.
About the study
In the present study, the researchers perform the first cell-type-specific integrative genomics study that includes three-dimensional (3D) genome mapping, HIV integration site sequencing (IS-seq), and transcription of a single core.
While previous studies used bulk tissue, the researchers in the current study used frontal lobe tissue from deceased HIV-infected donors and controls with and without encephalitis. More specifically, frontal cortex gray matter and subcortical white matter samples were obtained from three HIV-uninfected, three HIV-infected, and seven HIVE.
For transcript resolution at the cell-type level, the team performed 10X Chromium single-core ribonucleic acid sequencing (snRNA-seq) on tissue samples from the frontal lobe of the brain. In addition, the researchers examined single nuclei for HIV transcripts and observed HIV transcripts in all types of brain cells.
3D reorganization of microglia has not been visualized in vivo. The team therefore generated genome-scale high-throughput chromosome conformation capture (Hi-C) maps for microglia from two HIVE brains with age- and sex-matched controls. Based on sc RNA sequencing results, these HIVE brain samples had confirmed HIV expression.
In addition, the researchers examined 0.1 to 10 megabase (Mb) chromosomal compartments of microglia cells that were further divided into gene-poor and gene-poor A and B compartments. In particular, phase separation drives the spatial segregation of A and B chromosome compartments.
Because HIVE also remodels chromatin compartment architecture, the researchers explored topologically associated domains (TADs) and loop alterations in microglial cells.
Results of the study
Using snRNA-seq, 69,843 nuclei were profiled, resulting in an average of 2,401 differentially expressed genes (DEGs) and an average depth of 119,797 reads. In addition, analysis of differential Hi-C (dcHiC) compartments spanning 194 Mb of the microglial genome, which is nearly the size of human chromosome 5 in length, also revealed significant variations in A/B compartmentalization in the HIVE brain compared to the HIV-infected brain.
While undergoing profound reorganization, chromosomal conformations at the site of DEGs in microglia also altered kilo- to megabase-scale TADs and reconnected hundreds of contact-specific loops.
Compartment A genome of HIVE and HIV-infected microglia showed a marked change from 47.3% to 46.2%. Furthermore, 118 Mb of A/B chromosomal compartments in HIVE microglia cells shifted to a more open conformation, while 76 Mb transitioned to a more closed conformation in the opposite direction.
Regions with increased A compartmentalization in seven HIVE samples had 1,940 genes expressed at significantly higher levels than three HIV-infected microglia in the snRNA-seq dataset. In general, the changes of the Hi-C compartment in HIVE were specific to microglia.
Functional pathway analysis revealed substantial enrichment in complement cascade genes, interferon (IFN), and other cytokine signaling pathways, as well as myeloid chemotaxis and migration pathways. These changes pointed to a shift in microglia functionality from neuronal support functions to inflammation due to HIV integration in the brain.
Remarkably, non-encephalitic microglia from HIV-infected brains showed conserved changes in gene expression related to neuronal health, even in the absence of HIV integration. An early insult likely leads to HAND and associated neuronal dysfunction during HIV infection, despite the absence of direct neuronal infection.
Conclusions
The current study demonstrated that 3D genome-wide alterations in HIV-infected brains were microglia-specific and resulted in significant reprogramming of the nuclear transcriptome. For example, HIVE microglia gene loci, with upregulated expression, shifted to a more favorable transcriptional and open chromatin state, thus indicating that HIV integration and transcription caused these immune changes in microglia.
In addition, HIV-derived transcripts affected the chromosomal conformations of microglia through other mechanisms. HIV showed a strong predilection for integration into the open chromatin of compartment A. Indeed, some HIV integration sites likely depend on disease stages similar to T cells, which have been reported in previous studies.
Other viruses, such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), also rewire the chromosomal organization in the odorant receptor gene. The researchers observed a link between immune stimulation and infection, suggesting that activation of brain microglia potentiates HIV infection in some way.
These microglia are sites of high levels of productive HIV transcription, which then spreads from there to other susceptible brain cell populations. These small clonal populations of HIV integration sites reproduce and increase the CNS virus population.
Overall, data from the study showing cell type-specific differences in HIV integration sites could support the development of next-generation HIV cure therapies.
Journal reference:
- Plaza-Jennings, AL, Valada, A., O’Shea, C., et al. (2022). HIV integration into the human brain is linked to microglia activation and 3D genome remodeling. Molecular Cell. doi:10.1016/j.molcel.2022.11.016