Fig 2: FBN1 combines directly with VEGFR2 in ovarian cancer cells. (A) Co‐IP assay of interactions between FBN1 and VEGFR2 proteins in cisplatin‐resistant ovarian cancer organoids and OVCA433‐CisR cells. (B‐C) Interactions between FBN1 and VEGFR2 confirmed by FRET‐FLIM upon transient co‐expression in OVCA433‐CisR cells. **, P < 0.01. (D) Western blotting assay for determining the relationship between FBN1 and VEGFR2, p‐VEGFR2, downstream molecules of VEGFR2‐mediated signaling in ovarian cancer cells. (E) Immunofluorescence assay for determining the relationship between FBN1 and p‐VEGFR2 (Tyr1054) in cisplatin‐resistant ovarian cancer organoids and cell lines. Green signals, FBN1; red signals, p‐VEGFR2 (Tyr1054); blue signals, DAPI. (F) Western blotting assay of p‐AKT1 (Ser473) in cisplatin‐resistant ovarian cancer organoids and OVCA433‐CisR cells. (G) Co‐IP assay demonstrated an interaction between VEGFR2 domains 2 & 3 and FBN1 protein. Binding of VEGFA and the extracellular domains 2 & 3) of VEGFR2 was used as the positive control. (H) Mutant codons and amino acids of D2 and D3 of VEGFR2. (I) Western blotting assay of p‐AKT1 (Ser473) in cisplatin‐resistant ovarian cancer organoids and OVCA433‐CisR cells. Abbreviations: FBN1, fibrillin‐1; CR, cisplatin‐resistant; KO, knockout; NC, negative control; SDS‐PAGE, sodium dodecyl sulfate‐polyacrylamide gel electrophoresis; AKT, protein kinase B; Co‐IP, co‐Immunoprecipitation; VEGFR2, vascular endothelial growth factor receptor 2; DAPI, 2‐(4‐amidinophenyl)‐6‐indolecarbamidine dihydrochloride; FRET‐FLIM, Fӧrster resonance energy transfer‐fluorescence lifetime imaging. FE, FRET efficiency

Fig 3: STAT2 is the downstream target molecule of the FBN1/VEGFR2 signaling axis. (A) GSEA analysis was performed using FBN1‐knockout and control cisplatin‐resistant ovarian cancer organoids (CR‐organoids/FBN1‐KO1 and CR‐organoids/NC). The signature was defined by genes showing significant expression changes. (B) Effects of FBN1 knockout on mRNA levels of STAT family members. (C) FBN1 knockout greatly altered the distribution and expression of STAT2 and p‐STAT2 (Tyr690) in cytoplasm and nucleus of OVCA433‐CisR ovarian cancer cells. (D) Immunofluorescence assay to determine the association between p‐STAT2 (Tyr690) and FBN1 in cisplatin‐resistant ovarian cancer organoids and cells. (E) Western blotting assay to detect expression of STAT2 and p‐STAT2 (Tyr690) in cisplatin‐resistant ovarian cancer organoids and OVCA433‐CisR cells. The protein quantification was analyzed by Image J software. (F) mRNA expression of glycolysis and angiogenesis‐associated genes in FBN1‐knockout and control cisplatin‐resistant ovarian cancer organoids and OVCA433‐CisR cells. (G) Effect of FBN1 knockout, VEGFR2 overexpression, and STAT2 knockdown on HUVEC tube formation. HUVEC cells were treated with supernatant obtained from groups of NC, FBN1 KO‐1, FBN1 KO‐1+VEGFR2 OE, and FBN1 KO‐1+VEGFR2 OE+shSTAT2 in OVCA433‐CisR cells. Abbreviations: FBN1, fibrillin‐1; CR, cisplatin‐resistant; KO, knockout; NC, negative control; OE, overexpression; STAT, signal transducer and activator of transcription; VEGFR2, vascular endothelial growth factor receptor 2