Targeted Downregulation of TAZ Expression

In order to downregulate TAZ expression, we transfected cells with small interfering RNA (siRNA) designed to target human TAZ. The siRNA suspension was composed of Lipofectamine RNAiMAX (Thermo Fisher Scientific), OptiMEM medium (Thermo Fisher Scientific), and 20 nM of siRNA. Experiments were performed with three different siRNA that target TAZ: siTAZ1 (5′- ACGUUGACUUAGGAACUUU-3′, Eurofins, Dortmund, Germany) [47 (link)], siTAZ2 (5′-AGGUACCUUCCUCAAUACA-3′, Eurofins) [47 (link)], and siTAZ3 (5′-UGUGGAUGAGAUGGAUACA-3′, Eurofins). siTAZ1 and siTAZ2 are specific of TAZ sequences. siTAZ3 recognizes a common sequence of both TAZ and YAP mRNA. A negative control siCtrl (siCtrl 5′-GGGCAAGACGAGCGGGAAG-3′, Eurofins) with no known homology to mammalian genes was used. Two rounds of siRNA transfection were performed (separated by 24 h) in medium without antibiotics. Co-culture experiments were performed 6 h after the last transfection. Analyses were performed 30 h after the last transfection and 24 h after infection [13 (link),30 (link)]. TAZ inhibition efficiency was evaluated by Western blot experiments.