Interactions between the Sonic Hedgehog signaling pathway and the cAMP/PKA signaling pathway in LKB1 transfected breast cancer cells

Objective To investigate the interactions between the LKB1 tumor suppressor,the Sonic Hedgehog(SHH) signaling pathway and the cAMP/PKA signaling pathway.Methods The LKB1 gene was reintroduced into MDA-MB-231 breast cancer cells which were lack of LKB1.And then two groups were classified:MDA-MB-231 group(231 group) and LKB1 transfected MDA-MB-231 group(LKB1 group).The cells in each group were treated with the SHH signaling inhibitor cyclopamine at different concentration levels(0,5×10-6,1×10-5 and 2×10-5mol/L),respectively.The levels of SHH,Smo genes mRNA and protein expressions related to the SHH signaling pathway were detected with the methods of RT-PCR and Western blot analysis.Meanwhile,the activities of cAMP and PKA were determined with corresponding kits.Results After treated with different concentration of SHH signaling inhibitor cyclopamine,SHH,Smo genes mRNA and protein expressions related to the SHH signaling pathway in LKB1 group were significantly inhibited in comparison with those in 231 group.The inhibition effect was positively correlated to the concentration of cyclopamine,which reached the highest when the concentration of cyclopamine was 1×10-5mol/L.Above this concentration,the inhibition effect remained unaffected.Meanwhile,the values of PKA and cAMP in 231 group and LKB1 group both increased as cyclopamine concentration increased,and these values simultaneously achieved maxima when the concentration of cyclopamine was 1×10-5mol/L.Further increase of cyclopamine concentration to 2×10-5mol/L did not lead to any raise of PKA or cAMP values.Under the treatment with the same concentration of cyclopamine,higher values of PKA and cAMP were found in LKB1 group compared with 231 group.Conclusions In breast cancer MDA-MB-231 cells,the SHH signaling pathway is inhibited,under the synergetic affection of the LKB1 tumor suppressor and the cyclopamine inhibitor,while the expression of the cAMP/PKA signaling pathway is enhanced.When the concentration of cyclopamine was 1×10-5mol/L,the highest effect was observed.Therefore,it is supposed that there exists a LKB1-SHH-cAMP/PKA pathway.