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Autophagyhasbeenshowntopromoteinvasionofcancercellsduringstarvationorhypoxia.Wereportedthatautophagyacceleratedinvasionofhepatocarcinomacellsbyinducingepithelialmesenchymaltransition(EMT),whichchangedthecellphenotypefromepithelialintomesenchymalunderstarvationconditions.Transforminggrowthfactor(TGF)isakeycytokinethatinducesEMTinmanytypesofepithelialcells.Inourpreviousstudy,weshowedthatTGFexpressionandactivationofTGFsignallingplayedcrucialrolesinautophagyinducedEMTandinvasionofhepatocarcinomacells.However,themechanismunderlyingautophagicinductionofTGFEMTandinvasionofhepatocarcinomacellsisunclear.cAMPresponseelementbindingprotein(CREB)isaphosphoryla-dependenttranscriptionfactorthatisphosphorylatedbymulti-pleproteinkinasesandparticipatesindifferentproteinkinasesignaltransductionpathways.ActivationoftheCREBsignallingpathwayandphosphorylationofCREBwereshowntofulfillnumerouscellularfunctionsrangingfromcellproliferationandthecellcycletocelldif-ferentiationandcytokineproductionbybindingofphosphorylatedCREBtothecAMPresponseelement(CRE)intargetgenesandpro-motingtheirtranscription.ACREsitehasbeenidentifiedinthegenepromoter,andpCREBinducesTGFexpressioninbothnormalcellsandcancercellsbydirectlybindingtotheTGF FIGURE1AutophagycontributestodegradationofPDE4Ainhepatocarcinomacells.StarvationofHepG2andBEL7402cellsinHanksbalancedsaltsolution(HBSS)for24hinducedautophagy,whichwasshownbytheLC3ItoLC3conversionandP62degradation.InductionofautophagysignificantlyreducedPDE4Aexpressioninhepatocarcinomacells.TreatmentofcellsinHBSSwithchloroquine(5L)for24horcombinedtransfectionofcellswithsiRNAAtg3andsiRNAAtg7inhibitedautophagyandpreservedexpressionofPDE4Ainhepatocarcinomacells.A,RepresentativeWesternblotsand(B)densitometricanalysisofLC3I,LC3II,P62andPDE4AnormalizedtoactininHepG2(leftpanel)andBEL7402cells(rightpanel)culturedincompletemedium(CM)orinHBSSwithdifferenttreatments.CellsthatwereculturedinCMservedasacontrol.C,PDE4AdegradationkineticsatthemRNAlevelinHepG2(leftpanel)andBEL7402cells(rightpanel)thatwereculturedinCMandHBSSfor0,6,12,and24hweredeterminedbyquantitativeRTPCR.mRNAlevelsofPDE4Awerenormalizedtothatactin.D,RepresentativeWesternblotsand(E)densitometricanalysisforPDE4AdegradationkineticsnormalizedtoinHepG2(leftpanel)andBEL7402cells(rightpanel)thatwereculturedinCMandinHBSSfor0,6,12,and24h.CellsthatwereculturedinCMattheinitialtime(0h)servedasthecontrol.Dataarerepresentativeofthreeindependentexperimentsandareshownasthemean