Comparison of G4 abundance in TRRs of transcripts (the <i>F</i> score).

*<p>The <i>F</i>_cd, <i>F</i>_ncd, and <i>F</i>_o values from 1^st iteration were used as templates and compared with those from following iterations by one-way ANOVA test. No differences among iterations were observed (<i>P</i>≥0.05).</p

^MSN1DC₅₀ and ^{−80 G4}DC₅₀ values (µM) determined using FID assay for complexes 1–3 (values are average of three independent measurements and error is estimated to be below 5%).

<p>^MSN1DC₅₀ and ^{−80 G4}DC₅₀ values (µM) determined using FID assay for complexes 1–3 (values are average of three independent measurements and error is estimated to be below 5%).</p

CD spectra of the two promoter G-quadruplex sequences in solutions containing different salts.

<p>(<b>a</b>) single-repeat MNSG4. (<b>b</b>) −80 G4.</p

Transcription activity changes of human <i>TnIc</i> promoters in the presence of compound 3.

<p>(<b>a</b>) Transcription activity of constructed human <i>TnIc</i> promoters in the absence of compound <b>3</b>. (<b>b</b>) Transcription activity of constructed human <i>TnIc</i> promoters in the presence of 100 µM compound <b>3</b>. Columns in a and b are normalised to the luciferase transcription level of the wild type control 6MNS(E)-WT in the absence of ligand. (<b>c</b>) Dose-response curves of compound <b>3</b> with different luciferase vectors. (**<i>P</i><0.01, *<i>P</i><0.05 and ♯<i>P</i>≥0.05, no significant differences compared to WT construct).</p

Structure of human TnIc gene promoter and constructed luciferase expression vectors.

<p>(<b>a</b>) Schematic illustration of the G4-forming motifs in the promoter of human <i>TnIc</i> gene. (<b>b</b>) the structure of luciferase expression vectors driven by engineered human <i>TnIc</i> promoters.</p

Comparison of G4 location significance in TRRs of transcripts (the <i>Q</i> score).

*<p>The <i>Q</i>_cd, <i>Q</i>_ncd, and <i>Q</i>_o values from 1^st iteration were used as templates and compared with those from following iterations by one-way ANOVA test. No differences among iterations were observed (<i>P</i>>0.23).</p

TnIc G4s unfolding curves and corresponding fitted curves.

<p>(<b>a</b>) Decrease of Cy5 fluorescence in the process of MNSG4 (formed between oligos <b>Tr_MNS-III</b> and <b>Comp-Cy5</b>) opened by its C-rich complimentary strand (oligo <b>Tr_MNS-II</b>) at different temperature. Black, green, blue and cyan curves are from experimental data obtained at 25, 35, 45, and 55°C, respectively, and red curves are fitted results by a double exponential decay model. (<b>b</b>) Decrease of Cy5 fluorescence in the process of −80 G4 (formed between oligos <b>Tr_-80-III</b> and <b>Comp-Cy5</b>) opened by its C-rich complimentary strand (oligo <b>Tr_-80-II</b>) at different temperature. Black, green, blue and cyan curves are from experimental data obtained at 35, 45, 55, and 65°C, respectively, and red curves are fitted results by a double exponential decay model. Hybridisation was performed between 100 nM G4 forming oligos and 2 µM complimentary C-rich oligo. The reaction was carried out in 100 mM KCl solution containing 10 mM Tris-HCl (pH 7.4).</p

Structures and dose-response curves of compound 1, 2, and 3.

<p>(<b>a</b>) Compound <b>1</b> only slightly activated downstream luciferase expression (EC₅₀ = 812.1 µM). (<b>b</b>) Compounds <b>2</b> moderately activated downstream luciferase expression (EC₅₀ = 1.6 µM). (<b>c</b>) Compound <b>3</b> significantly activated downstream gene expression (EC₅₀ = 0.6 µM). Experiments were carried out by measuring the firefly (hTnIc-6MNS(E)-WT) and <i>Renilla</i> luciferase ratio changes in the presence of ligand with different concentrations.</p

Details of oligos used in biophysical studies in solution.

<p>Note: ^Cy3/5N indicates a Cy3 or Cy5-labeled nucleotide, which is coupled via NHS ester.</p

Energetics of TnIc MNSG4 and −80 G4 unfolding in solutions determined by CD melting.

<p>Experiments were carried out in solutions containing 10 mM Tris-HCl (pH 7.4) and 100 mM K⁺.</p