The impact of HTLV-1 expression on the 3D structure and expression of host chromatin.

A typical HTLV-1-infected individual carries >104 different HTLV-1-infected T cell clones, each with a single-copy provirus integrated in a unique genomic site. We previously showed that the HTLV-1 provirus causes aberrant transcription in the flanking host genome and, by binding the chromatin architectural protein CTCF, forms abnormal chromatin loops with the host genome. However, it remained unknown whether these effects were exerted simply by the presence of the provirus or were induced by its transcription. To answer this question, we sorted HTLV-1-infected T-cell clones into cells positive or negative for proviral plus-strand expression, and then quantified host and provirus transcription using RNA-seq, and chromatin looping using quantitative chromosome conformation capture (q4C), in each cell population. We found that proviral plus-strand transcription induces aberrant transcription and splicing in the flanking genome but suppresses aberrant chromatin loop formation with the nearby host chromatin. Reducing provirus-induced host transcription with an inhibitor of transcriptional elongation allows recovery of chromatin loops in the plus-strand-expressing population. We conclude that aberrant host transcription induced by proviral expression causes temporary, reversible disruption of chromatin looping in the vicinity of the provirus.

Time-course of host cell transcription during the HTLV-1 transcriptional burst.

The human T-cell leukemia virus type 1 (HTLV-1) transactivator protein Tax has pleiotropic functions in the host cell affecting cell-cycle regulation, DNA damage response pathways and apoptosis. These actions of Tax have been implicated in the persistence and pathogenesis of HTLV-1-infected cells. It is now known that tax expression occurs in transcriptional bursts of the proviral plus-strand, but the effects of the burst on host transcription are not fully understood. We carried out RNA sequencing of two naturally-infected T-cell clones transduced with a Tax-responsive Timer protein, which undergoes a time-dependent shift in fluorescence emission, to study transcriptional changes during successive phases of the HTLV-1 plus-strand burst. We found that the transcriptional regulation of genes involved in the NF-κB pathway, cell-cycle regulation, DNA damage response and apoptosis inhibition were immediate effects accompanying the plus-strand burst, and are limited to the duration of the burst. The results distinguish between the immediate and delayed effects of HTLV-1 reactivation on host transcription, and between clone-specific effects and those observed in both clones. The major transcriptional changes in the infected host T-cells observed here, including NF-κB, are transient, suggesting that these pathways are not persistently activated at high levels in HTLV-1-infected cells. The two clones diverged strongly in their expression of genes regulating the cell cycle. Up-regulation of senescence markers was a delayed effect of the proviral plus-strand burst and the up-regulation of some pro-apoptotic genes outlasted the burst. We found that activation of the aryl hydrocarbon receptor (AhR) pathway enhanced and prolonged the proviral burst, but did not increase the rate of reactivation. Our results also suggest that sustained plus-strand expression is detrimental to the survival of infected cells.