The pairs that are observed in a given number of optimal solutions for subtypes B and C.

<p>Listed for each of the optimal separating pairs are the covarying positions, the amino acids for the sequences in the Founder separating pairs, and then the Env motifs for each of these positions. The optimization problem was solved using objective i) (minimizing the number of pairs) on a covariance network constructed using all sequences in each subtype.</p

Comparison of optimal Founder pairs with the AA combinations appearing for 5 subtype B pre-seroconverters with each of the clones sequenced.

<p>Comparison of optimal Founder pairs with the AA combinations appearing for 5 subtype B pre-seroconverters with each of the clones sequenced.</p

Conserved and covarying regions for subtypes B and C <i>Env</i>.

<p>Regions are coloured as conserved across both subtypes (black), conserved within each subtype (dark blue), conserved except for a maximum of 2 individuals in that subtype (light blue), and covarying (magenta). Those covarying pairs with at least 20% of the maximum covariance value for subtype C, and 12.5% for subtype B, are connected with magenta lines. The different levels of covariance were determined to include approximately the same numbers of covarying pairs in each case: 78 for subtype B and 79 for subtype C. The signal, α₄β₇ binding site, constant (C1-C6) and variable (V1-V5) regions within gp120, and the gp41 domain are mapped onto the <i>Env</i> sequence.</p

Pairs observed multiple times (frequency f) in optimal networks for each subtype.

<p>The number of individuals exhibiting each AA combination is denoted by n.</p

Unrooted phylogenetic trees for the subtype B and C HIV Env sequences.

<p>Founder sequences are shown with red dashed branches, while chronic sequences are denoted by blue branches.</p

The collection of optimal pairs displayed relative to the domains of Env.

<p>A) over a linear representation and B) as networks.</p

Single AA in optimal networks determined on Founders, which appear at least 2 times (frequency f).

<p>The region of Env is denoted ahead of a decimal point, and any recognised motif after the decimal.</p

MOESM1 of HIV latency reversing agents act through Tat post translational modifications

Additional file 1: Figure S1. Cellular toxicity of LRAs. The CellTiter 96 Aqueous One Solution Cell Proliferation MTS assay was used to measure the toxicity of a panel of LRAs on HEK293T cells over a range of concentrations (31.25 to 1000 nM) for 48 h. VOR = vorinostat; PAN = panobinostat; CTN = chaetocin; DIS = disulfiram. The lines represent the mean + SD (n = 2

MOESM5 of HIV latency reversing agents act through Tat post translational modifications

Additional file 5: Table S1. Oligonucleotides used in this study fo

MOESM2 of HIV latency reversing agents act through Tat post translational modifications

Additional file 2: Figure S2. JQ1 increases EGFP and DsRed expression from an LTR-driven splicing reporter in the absence and presence of Tat. HEK293T cells were transfected with the pLTR.gp140/EGFP.Rev∆38/DsRed splicing reporter in the absence or presence of 100 ng of pTat101 (AD8)-Flag expression plasmid and then treated for 24 h with JQ1 (1 μM) or DMSO diluent control. Cells were harvested and portion analysed for either the percentage of cells expressing EGFP (unspliced, A.) or DsRed (spliced, B.) using flow cytometry, or HIV unspliced (US), spliced (D4-A7) and all viral RNA expression levels (copies/ul) by droplet digital PCR (ddPCR) (Fig. 3). The fold-change (FC) over DMSO of Live+ EGFP+ (A.), Live + DsRed + (B.) and percentage of spliced product DsRed/(DsRed + EGFP) (C.) were determined. Comparisons of each condition to DMSO were made using a paired T test. Only statistically significant comparisons are shown **p < 0.01; ***p < 0.001; ****p < 0.0001. The black lines represent the mean ± SEM (n = 4