Sitran sirna transfection reagent

Mammalian expression constructs of p300, YAP, TAZ, TEAD3, and TEAD4 (WT and the K297A, Y429A, and W299A mutants), fused with HA or Flag tag, were generated based on the pcDNA3.1(+) vector. The luciferase reporters, including CAGA-luciferase, ARE-luciferase, 3TP-luciferase, Gal4-luciferase (pFR-luciferase), and Renilla-luciferase, and constructs encoding Gal4-DBD, Gal4-Smad3, Flag-Smad3, Flag-Smad4, 6Myc-Smad2, 6Myc-Smad3, and 6Myc-Smad4 were described previously (Yan et al., 2016 (link), 2018 (link)).
Double stranded siRNAs targeting human YAP, TAZ, or TEAD4 and the non-specific control siRNA were purchased from RiboBio (China). siRNAs were transfected using the siTran^TM siRNA Transfection reagent (OriGene). The sequences of siRNAs used in this study were as follows: YAP siRNA, 5′-CCACCAAGCTAGAT AAAGA-3′; TAZ siRNA, 5′-CCGCAGGGCTCATGAGTAT-3′; TEAD4 siRNA #1: 5′-GTATGCTCGCTATGAGAAT-3′; and TEAD4 siRNA #2: 5′-GGAACAAACUGUGCCUGAATT-3′. Recombinant human TGF-β1 peptide was purchased from R&D Systems. In this study, TGF-β treatments all refer to recombinant TGF-β1 peptide.

Mammalian expression plasmids for Smad2, Smad3, Smad4, TEAD4, and YAP (WT and point mutants including 5SA, S94A, and 5SA/S94A) were fused with HA-, Flag-, or Myc-tags and constructed using the vector pcDNA3.1(+) or pCMV5 (53 (link), 54 (link), 55 ). The full-length complementary DNA of human CYR61 gene was inserted into the pcDNA3.1(+) with a C-terminal Flag tag. The CYR61 gene promoter regions, i.e., the −2475 to +15 bp DNA fragment from the TSS or different truncations of this region, were cloned into pGL3.0-basic vector to generate CYR61-pro-luciferase constructs.
The nonspecific control siRNA and those targeting human YAP, TAZ, or CYR61 were purchased from RiboBio. siRNAs were transfected with the siTran siRNA Transfection reagent (OriGene). The sequences of siRNAs used in this study were documented in Table S2.
Recombinant human TGF-β1 (CA59) and CYR61 (CB98) peptides were purchased from Novoprotein. Small chemical inhibitors, such as JSH-23 (HY-13982), SB216763 (HY-12012), verteporfin (HY-B0146), SB431542 (HY-10431), PD0325901 (HY-131295), SP600125 (HY-12041), LY3214996 (HY-101494), and LY294002 (HY-10108), were all purchased from MedChemExpress (MCE), while XMU-MP-1 was obtained from AbMole (M9057).