Braco 19

BRACO-19 hydrochloride (BRACO-19) was purchased from Sigma-Aldrich (#SML0560-5MG). TMPyP4 was purchased from Calbiochem-EMD Millipore (#613560-25MG). 0.5 × 10⁶ HEK293T cells were plated in 6-well plates and left untreated (control) or treated for 24 h with either BRACO-19 (0, 1, 3, and 32 µM) or TMPyP4 (0, 0.5, 1, and 8 µM). The BRACO-19 and TMPyP4 concentrations were established from previously used concentrations [54 (link),55 (link),56 (link)]. The cells were then infected for 5 h with pseudotyped HIV-1/VSV-G in the presence of 10 µg/mL of polybrene (Sigma-Aldrich, #H9268-5G). The medium was changed 5 h post-infection and the cells were treated anew with either BRACO-19 or TMPyP4 for 24 h. The genomic DNA was extracted from the cells as per the manufacturer’s instructions using the DNeasy Blood and Tissue Kit (Qiagen, Hilden, Germany, #69504) and processed for the integration site profile analyses.

Netropsin, BRACO-19, and pyridostatin were purchased from Sigma Aldrich. Phen-DC₃ was a gracious gift from Marie-Paule Teulade-Fichou. Lyophilized Phen-DC₃ was dissolved in dimethyl sulfoxide. All other ligands in lyophilized form were dissolved in water.

A subcohort of 2-month-old Atrx^HET and Atrx^HOM mice was administered either BRACO-19 (SML0560, Sigma-Aldrich) (2 mg/kg, SID, Monday to Friday, diluted in distilled water (dH₂O)) or vehicle (dH₂O) via IP, for 20 or 40 days, as previously performed [43 (link)]. The detailed composition of each treatment group can be consulted in Supplementary File S1: Table S4. Besides weight analysis, the experimental unit of this experience outcome was not the individual mice but the acquired images of pancreatic islets.

DNA oligonucleotides were purchased from TIBMOLBIOL (Berlin, Germany) and further purified through ethanol precipitation prior to their use. Concentrations of oligonucleotides were determined by measuring their absorbance A₂₆₀ at 80°C in aqueous solution using extinction coefficients as provided by the manufacturer. PIQ derivatives were prepared as described and their concentration determined spectrophotometrically using a molar extinction coefficient ϵ₃₇₆ of 22 227 M⁻¹·cm⁻¹ in potassium phosphate buffer (28 (link),29 (link)). Thiazole orange (TO), SYUIQ-5, BRACO-19 and Phen-DC3 were purchased from Sigma-Aldrich Chemie GmbH (Taufkirchen, Germany). NDI-DM was obtained from ABCR (Karlsruhe, Germany). Thiazole orange concentrations were determined using a molar extinction coefficient for the TO monomer ϵ₅₀₀ of 63 000 M⁻¹·cm⁻¹ in DMSO (30 (link)). Concentrations of all other ligands were determined from their weighed mass. All ligands were initially dissolved in a DMSO stock solution except for isothermal titration calorimetry (ITC) measurements. Here, ligands were directly dissolved in the high-salt ITC buffer solution (20 mM potassium phosphate, 100 mM KCl, pH 7.0, supplemented with 5% DMSO). A low-salt buffer with 10 mM potassium phosphate, pH 7.0, was additionally employed for circular dichroism (CD) melting and some of the NMR experiments as described.

NMM and TMPyP2 were purchased from Frontier Scientific. TMPyP4 tosylate was bought from Abcam. BRACO19 and Pyridostatin (PDS) were purchased from Sigma-Aldrich.

Oligonucleotides used in CD analysis (Table 1) were obtained from Eurofins Genomics (Ebersberg, Germany) (https://www.eurofinsgenomics.eu/). BRACO-19 and pyridostatin were obtained from Merck-Sigma (Milan, Italy). Stock solutions were prepared in H₂O at 1 mM and further diluted for subsequent experiments.