Biology subjects

Svensson, J. P.

Publications and source records attributed to Svensson, J. P..

5 recordsLinked to original sources

bioRxiv2024

De novo PADI4-mediated citrullination of histone H3 stimulates HIV-1 transcription

HIV-1 infection establishes a reservoir of long-lived cells with integrated proviral DNA that can persist despite antiretroviral therapy (ART). Some of these reservoir cells can at anytime be reactivated and reinitiate infection. The mechanisms governing proviral latency the transcriptional regulation of the provirus are complex and have not yet been sufficiently described. Here, we identified a role for histone H3 citrullination, a post-translational modification catalyzed by protein-arginine deiminase type-4 (PADI4), in HIV-1 transcription and latency. We found that PADI4 inhibition by GSK484 reduced HIV-1 transcription after T cell activation in ex vivo cultures of CD4 T cells from people living with HIV-1 (PLWH). The effect was more pronounced in individuals with active viremia compared to individuals with effective ART. Using cell models of HIV-1 latency, we showed that PADI4-mediated citrullination of histone H3 occurred at the HIV-1 promoter upon T cell stimulation which facilitated proviral transcription. HIV-1 preferentially integrated into genomic regions marked by H3 citrullination and these integrated proviruses were less prone to latency compared to those in non-citrullinated chromatin. Inhibiting PADI4 led to compaction of the HIV-1 promoter chromatin and an increase of HP1-covered heterochromatin, in a mechanism partly dependent on the HUSH complex. Our data reveal a novel mechanism to explain HIV-1 latency and transcriptional regulation. HighlightsO_LIThe PADI4 enzyme stimulates HIV-1 transcription during T cell activation. C_LIO_LIPADI4 citrullinates histone H3 at the HIV-1 promoter upon T cell activation and inhibiting PADI4 reduces HIV-1 reactivation in ex vivo CD4 T cells from people living with HIV-1. C_LIO_LIH3cit is mostly found at gene promoters, and productive HIV-1 proviruses are more likely than latent or reactivatable proviruses, to integrate in chromatin susceptible for citrullination. C_LIO_LIH3cit inhibits latency establishment by interfering with the binding of HP1 to H3K9me3, preventing heterochromatin formation. C_LI

microbiology↗

bioRxiv2021

T cell stimulation remodels the latently HIV-1 infected cell population by differential activation of proviral chromatin

The reservoir of latently HIV-1 infected cells is heterogeneous. To achieve an HIV-1 cure, the reservoir of activatable proviruses should be eliminated while permanently silenced proviruses may be tolerated. We have developed a method to assess the proviral nuclear microenvironment in single cells. In latently HIV-1 infected cells, a zinc finger protein tethered to the HIV-1 promoter produced a fluorescent signal as a protein of interest came in its proximity, such as the viral transactivator Tat when recruited to the nascent RNA. Tat is essential for viral replication. In these cells we assessed the proviral activation and chromatin composition. By linking Tat recruitment to proviral activity, we dissected the mechanisms of HIV-1 latency reversal and the consequences of HIV-1 production. A pulse of promoter-associated Tat was identified that contrasted to the continuous production of viral proteins. As expected, promoter H3K4me3 led to substantial expression of the provirus following T cell stimulation. However, the activation-induced cell cycle arrest and death led to a surviving cell fraction with proviruses encapsulated in repressive chromatin. Further, this cellular model was used to reveal mechanisms of action of small molecules. In a proof-of-concept study we determined the effect of an enhancer specific CBP/P300-inhibitor on HIV-1 latency reversal. Only proviruses resembling active enhancers, associated with H3K4me1 and H3K27ac, efficiently recruited Tat. Tat-independent HIV-1 latency reversal of unknown significance still occurred. We present a method for single cell assessment of the microenvironment of the latent HIV-1 proviruses, used here to reveal how T cell stimulation modulates the proviral activity and how the subsequent fate of the infected cell depends on the chromatin context.

microbiology↗

bioRxiv2019

Chromatin maturation of the HIV-1 provirus in primary resting CD4+ T cells

Human immunodeficiency virus type 1 (HIV-1) infection is a chronic condition, where viral DNA integrates into the genome. Latently infected cells form a persistent, heterogeneous reservoir. The reservoir that reinstates an active replication comprises only cells with intact provirus that can be reactivated. We confirmed that latently infected cells from patients exhibited active transcription throughout the provirus. To find transcriptional determinants, we characterized the establishment and maintenance of viral latency during proviral chromatin maturation in cultures of primary CD4+ T-cells for four months after ex vivo HIV-1 infection. As heterochromatin (marked with H3K9me3 or H3K27me3) gradually stabilized, the provirus became less accessible with reduced activation potential. In a subset of infected cells, active marks (i.e., H3K27ac) remained detectable, even after prolonged proviral silencing. After T-cell activation, the proviral activation occurred uniquely in cells with H3K27ac-marked proviruses. Our observations suggested that, after transient proviral activation, cells were actively returned to latency.

microbiology↗

bioRxiv2018

The ultra-sensitive Nodewalk technique identifies stochastic from virtual, population-based enhancer hubs regulating MYC in 3D: Implications for the fitness of cancer cells

The relationship between stochastic transcriptional bursts and dynamic 3D chromatin states is not well understood due to poor sensitivity and/or resolution of current chromatin structure-based assays. Consequently, it is not well established if enhancers operate individually and/or in clusters to coordinate gene transcription. In the current study, we introduce Nodewalk, which uniquely combines high sensitivity with high resolution to enable the analysis of chromatin networks in minute input material. The >10,000-fold increase in sensitivity over other many-to-all competing methods uncovered that active chromatin hubs identified in large input material, corresponding to 10 000 cells, flanking the MYC locus are primarily virtual. Thus, the close agreement between chromatin interactomes generated from aliquots corresponding to less than 10 cells with randomly re-sampled interactomes, we find that numerous distal enhancers positioned within flanking topologically associating domains (TADs) converge on MYC in largely mutually exclusive manners. Moreover, when comparing with several enhancer baits, the assignment of the MYC locus as the node with the highest dynamic importance index, indicates that it is MYC targeting its enhancers, rather than vice versa. Dynamic changes in the configuration of the boundary between TADs flanking MYC underlie numerous stochastic encounters with a diverse set of enhancers to depict the plasticity of its transcriptional regulation. Such an arrangement might increase the fitness of the cancer cell by increasing the probability of MYC transcription in response to a wide range of environmental cues encountered by the cell during the neoplastic process.

genomics↗

bioRxiv2018

Chromatin Functional States Correlate with HIV Latency Reversal in Infected Primary CD4+ T Cells

Human immunodeficiency virus (HIV) infection cannot be cured due to a small reservoir of latently infected CD4+ T cells in treated patients. The \"shock and kill\" approach proposes to eliminate the reservoir by inducing its activation and the direct or indirect killing of infected cells. Current latency reversing agents (LRAs) do not reduce the viral reservoir in treated patients.\n\nWe use a novel dual-fluorescent HIV reporter to identify and purify latent cells, and to determine the fraction of latent cells that undergo viral reactivation after infection of primary CD4+ T cells. Unexpectedly, LRAs reactivate less than 5% of latent proviruses. Analysis of HIV integration sites from induced and non-induced latent populations reveals distinct provirus integration sites between these two populations in terms of chromatin functional states.\n\nThese findings challenge \"shock and kill\", and suggest the need of more potent LRAs in combination with immunomodulatory approaches to eradicate HIV reservoir.

microbiology↗