Figure 6. Energy stress attenuates tumorigenecity of Lats1/2 DKO MEFs.

(a) AICAR inhibits YAP target gene expression in Lats1/2 DKO MEFs. Lats1/2 DKO MEFs were treated with 0.5 mM and 1 mM of AICAR for 5 hr. mRNA levels of Ctgf and Cyr61 were measured by quantitative RT-PCR (error bars represent ± s.e.m. from n=3 independent experiments). (b) Knockdown of YAP or TAZ attenuates anchorage-independent growth of Lats1/2 DKO MEFs. YAP or TAZ was knockdown by shRNAs in Lats1/2 DKO MEFs (the experiments was repeated 2 times). The Lats1/2 DKO MEFs (3 × 10³ cells) were seeded onto a culture medium containing 0.3% agar and incubated at 37 °C for 2 weeks. The colonies were stained with 0.005% crystal violet. (c) Energy stress attenuates anchorage-independent growth of Lats1/2 DKO MEFs. Lats1/2 DKO MEFs (3 × 10³ cells) were seeded onto a culture medium containing 0.3% agar and incubated at 37 °C for 2 weeks. The colonies were stained with 0.005% crystal violet and quantified. AICAR and metformin treatments were indicated (error bars represent ± s.e.m. from n=3 independent experiments). (d) Metformin inhibits tumor growth of Lats1/2 DKO MEFs in xenografted mice. Mice were ectopically implanted with 1x10⁶ cells on the both sides of each mouse. Daily i.p. injection of 250mg/kg dose of Metformin or equal volume (300µL) of vehicle (PBS) for 15 days. At the end of the experiment, mice were sacrificed 16 days after implantation and tumor weight was measured (error bars represent ± s.e.m. from 8 tumors (4 mice) per group). Scale bar, 1cm. ^*P<0.05, two-tailed t-test.