Hippo/MST blocks breast cancer by downregulating WBP2 oncogene expression via miRNA processor Dicer

摘要

Overexpression of MST and/or LATS kinase in combination with their regulatory proteins SAV and MOB abolished the WNT3A-induced, WBP2-mediated TCF reporter activity in HeLa. HeLa cells were transiently transfected with pCDNA6.2-WBP2 in various combinations with pCI-Neo-MST1, MST2, LATS1, LAST2, SAV, and MOB and TOPFlash luciferase reporter plasmids. Empty vector pCDNA6.2 or pCI-Neo was used as the negative control. Thirty-six hours after transfection, the cells were stimulated with L control-(O) and WNT3A-conditioned medium for 8h, followed by dual luciferase assay. QC of the Hippo components (MST1, MST2, LATS1, LATS2, SAV, MOB) overexpression and their activation in HeLa using immunoblotting. : Overexpression of Hippo pathway cassettes downregulated WBP2 protein expression in a dose-dependent manner in HeLa. HeLa cells were transiently transfected with pCDNA6.2-WBP2 in various combinations with pCI-Neo-MST1, MST2, LATS1, LAST2, SAV and MOB in the decreasing dose. Twenty-four hours after transfection, the total cell lysates were examined by western blot. (i) LATS1/2 knockdown did not abolish MST1/2-mediated WBP2 downregulation in HeLa. HeLa cells were transiently transfected with pCDNA6.2-WBP2 together with vector or pCI-Neo-MST1, MST2, and SAV in the presence luciferase or LATS1+2 siRNAs. Twenty-four hours after transfection, the total cell lysates were examined by western blot. (ii) LATS1/2-mediated WBP2 downregulation was independent of MST1/2 knockdown in HeLa. HeLa cells were transiently transfected with pCDNA6.2-WBP2 together with vector or pCI-Neo-LATS1, LATS2 and MOB in the presence luciferase or MST1+2 siRNAs. Twenty-four hours after transfection, the total cell lysates were examined by western blot. SAV potentiates MST1 and/or MST2-mediated downregulation of WBP2 in HeLa. HeLa cells were transiently transfected with pCDNA6.2-WBP2 together with MST1, MST2 and SAV individually and in combination. Twenty-four hours after transfection, the total cell lysates were examined by western blot.