Fig. 8

Fig. 8

CHEK2 activates autophagy to promote primary ciliation and cell invasion under serum starvation. A-C AMPK and Beclin 1 were activated during serum starvation. Extracts of HTR-8 cells serum-starved for 1 or 2 days were analyzed by Western blotting with antibodies against phosphorylated AMPK (p-AMPK), AMPK, phosphorylated ULK1, ULK1, phosphorylated Beclin1 (p-Beclin1), Beclin1, Ku70, and actin. B-C Quantitative results for the relative intensity of (B) p-AMPK/AMPK and (C) p-Beclin1/Beclin1 in A. D-F Inhibition of CHEK2 alleviated AMPK and Beclin1 activation. Extracts of HTR-8 cells in the absence or presence of CHEK2 inhibitor (CHEK2i) with or without serum starvation were analyzed by Western blotting with antibodies against phosphorylated AMPK (p-AMPK), AMPK, phosphorylated Beclin1 (p-Beclin1), Beclin1, Ku70, and actin. E-F Quantitative results for the relative intensity of (E) p-AMPK/AMPK and (F) p-Beclin1/Beclin1 in D. G-J Inhibition of AMPK reduced primary cilia and trophoblast cell invasion. G AMPK was inhibited by treating cells with AMPK inhibitor. Extracts of serum-starved HTR-8 cells in the presence or absence of AMPK inhibitor (AMPKi) were analyzed by Western blotting with antibodies against phosphorylated AMPK (p-AMPK), AMPK, and Ku70. H Quantitative results for the relative intensity of p-AMPK/AMPK in G. I-J Inhibition of AMPK reduced primary cilia and cell invasion HTR-8 cells. Quantitative results of (I) ciliated and (J) invaded cells in the absence or presence of AMPKi. K-N Depletion of Beclin1 reduced primary cilia and invasion of trophoblast cells. K-L Beclin1 was depleted efficiently. K Extracts of HTR-8 cells transfected with siRNA against Beclin1 were analyzed by Western blotting with antibodies against Beclin1 and actin. L Quantitative results for the relative intensity of p-Beclin1/Beclin1 in (K). (M-N) Quantitative results of (M) ciliated and (N) invaded cells of wild-type or Beclin1-depleted (siBeclin1) cells. O-P Inhibition of AMPK alleviated Beclin1 activation. O Extracts of cells treated with or without AMPK inhibitor (AMPKi) were analyzed by Western blotting with antibodies against phosphorylated Beclin1 (p-Beclin1), Beclin1, and Ku70. P Quantitative results for the relative intensity of p-Beclin1/Beclin1 in O. ***p < 0.001. These results are mean ± SD from three independent experiments