Alpha sma

After the ring was removed, the CAM area within a silicon ring was fixed on day 13 in formaldehyde for 24 h, which was followed by paraffin embedding as previously described [48 (link)]. Then, sections of five micrometer were cut tangentially to the CAM surface. Afterwards, for further histopathologic assessment of the vessels, a hematoxylin and eosin stain (H&E, Merck, Darmstadt, Germany), α-smooth muscle actin stain (alphaSMA, Sigma-Aldrich, St. Louis, MO, USA) and CD105 (Biorbyt, Cambridge, England) antibody stain were performed according to manufacturer’s instruction as described before [9 (link)] (Figure 1 and Figure 2). Finally, pictures of the slices (2× magnification) were taken with the light microscopy (KEYENCE, Neu-Isenburg, Germany) and analysed via the BZ-II Analyzer (KEYENCE, Neu-Isenburg, Germany) software for average area of vessel staining (average size of stained area, pixels) and total area (summarized amount of stained area, pixels) by H&E and alphaSMA slices. In case of H&E slices, an additionally brightness integration (sum of the whole brightness intensity of all ROIs, quadratcount method, pixels) was counted (Figure 2). The total area of the slices with corresponding values was equalized to 100% of one standardized area to compare the groups.

Standard western blotting techniques and Amersham ECL Plus Western blotting reagent were used. Following antibodies were used: JMJD1a (12835-1-AP, Proteintech, USA, 1:1,000), YAP/TAZ (sc-101199, Santa Cruz Biotechnology, USA, 1:500), GAPDH (5G4, HyB test 1:5,000), Tubulin (12G10, Hybridoma bank, 1:5,000), Lamin A/C (sc-7292, Santa Cruz Biotechnology, 1:1,000), H3K9me2 (#7658, Cell Signaling, 1:1,000), Histone 3 (#4499, Cell Signaling, 1:1,000), actin (clone AC-74, Sigma, 1:1,000), alpha-SMA (A2547, Sigma, 1:1,000) and antiphosphotyrosine antibody (APY03, Cytoskeleton, 1:1,000). For the phosphotyrosine pull-downs, MDA-MB-231 cells co-transfected with GFP-JMJD1a and CA-Src or empty vector were lysed and subjected to pulldown with beads of APY03 covalently coupled to sepharose (Anti-Phosphotyrosine Affinity Beads # APY03-Beads (Cytoskeleton) according to the manufacturer's instructions. The pulldowns and cell lysate were resolved on SDS–PAGE and subjected to western blot analysis with anti-GFP antibody (Abcam #1218).
Uncropped scans of the most important blots are provided as Supplementary Fig. 10.

Arsenic trioxide (Sigma-Aldrich) was prepared in 1 N NaOH at 250 mM and then diluted in sterile water for a stock concentration of 1 μM. Cell culture medium, FGM-2 and DMEM, were purchased from Lonza (Allendale, NJ) and Gibco. Human recombinant TGF-β1 was purchased from R&D systems (Minneapolis, MN). Antibodies used were: alpha-SMA (Sigma-Aldrich, 1:10,000), type-1 collagen (abcam, 1:2000), PML (Santa Cruz, 1:500), PAI-1 (peprotech, 1:2000), fibronectin (BD science, 1:500). Antibodies for Smad2, p-Samd2, Smad3, p-Smad3, Akt, p-Akt, Erk, p-Erk, p38, p-p38 were purchased from Cell signaling and used at a concentration of 1:1000.

We utilized the following antibodies: STING (catalog 13647 for WB, 67733 for IHC, CST), pSTING (catalog 50907, CST), pTBK1 (catalog 5483, CST), TBK1 (catalog 38066, CST), pIRF3 (catalog 37829, CST), IRF3 (catalog 4302, CST), alpha-SMA (catalog A2547, Sigma-Aldrich), Collagen type I (ab292, Abcam), Vimentin (catalog ab92547, Abcam), and actin (catalog AC-40, Sigma-Aldrich). Bindarit (catalog SML2910) was purchased from Sigma-Aldrich.

For immunofluorescence assays, frozen tissue or embryo sections were fixed, blocked and incubated with primary antibodies against PECAM-1, Wt1, Dystrophin and β -catenin, Gata5 (Santa Cruz), alpha-SMA (Sigma-Aldrich), PKR1 (IGBMC, Illkirch), GFP (Abcam), Myosin heavy chain epitope (MF20) (DSHB, University of Iowa), Akt (Cell signaling) and active-caspase-3 (Abcam). Phalloidin-488, zonula occluden-1 (ZO-1) (Invitrogen) was used to label F-actin to delineate the cellular cytoskeleton. Antibody binding was detected by incubation with Fluorescein, Alexa 555-, Alexa 488- or Alexa 594-conjugated (Millipore) secondary antibodies or Vectastain ABC peroxidase kit, according to the manufacturer’s instructions.