Skip to main content
. 2013 Jun 1;27(11):1223–1232. doi: 10.1101/gad.219402.113

Figure 3.

Figure 3.

PKA increases YAP phosphorylation by stimulating kinase activity of Lats1/2. (A) Myc-tagged wild-type or S127A or 5SA mutant YAP were transfected into HEK293A cells, and, after 16 h, cells were treated with or without 10 μM forskolin for 1 h. YAP phosphorylation was assessed by phos-tag gel. (B) MDA-MB-231 cells were transfected with control, MST1/2, or Lats1/2 siRNAs. Two days later, cells were treated with 10 μM epinephrine or forskolin for 1 h. Cell lysates were subjected to immunoblotting to assess knockdown efficiency and YAP phosphorylation. The arrowhead indicates MST2 position. (C,D) Flag-YAP was cotransfected into HEK293A cells with or without K/R mutants (kinase-dead) of MST2 (C) or Lats2 (D); after 16 h, cells were stimulated with 10 μM forskolin for 1 h, and phos-tag gels were used to determine the phosphorylation status of Flag-YAP. (E) MDA-MB-231 cells were untreated or treated with 10 μM forskolin for 1 h, endogenous Lats1 was immunoprecipitated and subjected to kinase assay using GST-YAP as substrate, and phosphorylation of GST-YAP by Lats1 was monitored by YAP phosphorylation at S127.