Genomic Exploration of the Brain in People Infected with HIV-Recent Progress and the Road Ahead.

PURPOSE OF REVIEW: The adult human brain harbors billions of microglia and other myeloid and lymphoid cells highly susceptible to HIV infection and retroviral insertion into the nuclear DNA. HIV infection of the brain is important because the brain is a potentially large reservoir site that may be a barrier to HIV cure strategies and because infection can lead to the development of HIV-associated neurocognitive disorder. To better understand both the central nervous system (CNS) reservoir and how it can cause neurologic dysfunction, novel genomic, epigenomic, transcriptomic, and proteomic approaches need to be employed. Several characteristics of the reservoir are important to learn, including where the virus integrates, whether integrated proviruses are intact or defective, whether integrated proviruses can be reactivated from a latent state to seed ongoing infection, and how this all impacts brain function. RECENT FINDINGS: Here, we discuss similarities and differences of viral integration sites between brain and blood and discuss evidence for and against the hypothesis that in the absence of susceptible T-lymphocytes in the periphery, the virus housing in the infected brain is not able to sustain a systemic infection. Moreover, microglia from HIV + brains across a wide range of disease severity appear to share one type of common alteration, which is defined by downregulated expression, and repressive chromosomal compartmentalization, for microglial genes regulating synaptic connectivity. Therefore, viral infection of the brain, including in immunocompetent cases with near-normal levels of CD4 blood lymphocytes, could be associated with an early disruption in microglia-dependent neuronal support functions, contributing to cognitive and neurological deficits in people living with HIV.

R-loop landscapes in the developing human brain are linked to neural differentiation and cell-type specific transcription.

Here, we construct genome-scale maps for R-loops, three-stranded nucleic acid structures comprised of a DNA/RNA hybrid and a displaced single strand of DNA, in the proliferative and differentiated zones of the human prenatal brain. We show that R-loops are abundant in the progenitor-rich germinal matrix, with preferential formation at promoters slated for upregulated expression at later stages of differentiation, including numerous neurodevelopmental risk genes. RNase H1-mediated contraction of the genomic R-loop space in neural progenitors shifted differentiation toward the neuronal lineage and was associated with transcriptomic alterations and defective functional and structural neuronal connectivity in vivo and in vitro. Therefore, R-loops are important for fine-tuning differentiation-sensitive gene expression programs of neural progenitor cells.

HIV integration in the human brain is linked to microglial activation and 3D genome remodeling.

To explore genome organization and function in the HIV-infected brain, we applied single-nuclei transcriptomics, cell-type-specific chromosomal conformation mapping, and viral integration site sequencing (IS-seq) to frontal cortex from individuals with encephalitis (HIVE) and without (HIV+). Derepressive changes in 3D genomic compartment structures in HIVE microglia were linked to the transcriptional activation of interferon (IFN) signaling and cell migratory pathways, while transcriptional downregulation and repressive compartmentalization of neuronal health and signaling genes occurred in both HIVE and HIV+ microglia. IS-seq recovered 1,221 brain integration sites showing distinct genomic patterns compared with peripheral lymphocytes, with enrichment for sequences newly mobilized into a permissive chromatin environment after infection. Viral transcription occurred in a subset of highly activated microglia comprising 0.33% of all nuclei in HIVE brain. Our findings point to disrupted microglia-neuronal interactions in HIV and link retroviral integration to remodeling of the microglial 3D genome during infection.

3D Genome Plasticity in Normal and Diseased Neurodevelopment.

Non-random spatial organization of the chromosomal material inside the nuclei of brain cells emerges as an important regulatory layer of genome organization and function in health and disease. Here, we discuss how integrative approaches assessing chromatin in context of the 3D genome is providing new insights into normal and diseased neurodevelopment. Studies in primate (incl. human) and rodent brain have confirmed that chromosomal organization in neurons and glia undergoes highly dynamic changes during pre- and early postnatal development, with potential for plasticity across a much wider age window. For example, neuronal 3D genomes from juvenile and adult cerebral cortex and hippocampus undergo chromosomal conformation changes at hundreds of loci in the context of learning and environmental enrichment, viral infection, and neuroinflammation. Furthermore, locus-specific structural DNA variations, such as micro-deletions, duplications, repeat expansions, and retroelement insertions carry the potential to disrupt the broader epigenomic and transcriptional landscape far beyond the boundaries of the site-specific variation, highlighting the critical importance of long-range intra- and inter-chromosomal contacts for neuronal and glial function.

Nanos-mediated repression of hid protects larval sensory neurons after a global switch in sensitivity to apoptotic signals.

Dendritic arbor morphology is a key determinant of neuronal function. Once established, dendrite branching patterns must be maintained as the animal develops to ensure receptive field coverage. The translational repressors Nanos (Nos) and Pumilio (Pum) are required to maintain dendrite growth and branching of Drosophila larval class IV dendritic arborization (da) neurons, but their specific regulatory role remains unknown. We show that Nos-Pum-mediated repression of the pro-apoptotic gene head involution defective (hid) is required to maintain a balance of dendritic growth and retraction in class IV da neurons and that upregulation of hid results in decreased branching because of an increase in caspase activity. The temporal requirement for nos correlates with an ecdysone-triggered switch in sensitivity to apoptotic stimuli that occurs during the mid-L3 transition. We find that hid is required during pupariation for caspase-dependent pruning of class IV da neurons and that Nos and Pum delay pruning. Together, these results suggest that Nos and Pum provide a crucial neuroprotective regulatory layer to ensure that neurons behave appropriately in response to developmental cues.