Although　PKC-mediated　phosphorylation　of　protein　kinase　D1　(PKD1)　has　been　extensively　characterized,　little　is　known　about　PKD1　regulation　by　other　upstream　kinases.　Here　we　report　that　stimulation　of　epithelial　or　fibroblastic　cells　with　G　protein-coupled　receptor　agonists,　including　angiotensin　II　or　bombesin,　induced　rapid　and　persistent　PKD1　phosphorylation　at　Ser(203),　a　highly　conserved　residue　located　within　the　PKD1　N-terminal　domain.　Exposure　to　PKD　or　PKC　family　inhibitors　did　not　prevent　PKD1　phosphorylation　at　Ser(203),　indicating　that　it　is　not　mediated　by　autophosphorylation.　In　contrast,　several　lines　of　evidence　indicated　that　the　phosphorylation　of　PKD1　at　Ser(203)　is　mediated　by　kinases　of　the　class　I　PAK　subfamily,　specifically　1)　exposing　cells　to　four　structurally　unrelated　PAK　inhibitors　(PF-3758309,　FRAX486,　FRAX597,　and　IPA-3)　that　act　via　different　mechanisms　abrogated　PKD1　phosphorylation　at　Ser(203),　2)　siRNA-mediated　knockdown　of　PAK1　and　PAK2　in　IEC-18　and　Swiss　3T3　cells　blunted　PKD1　phosphorylation　at　Ser(203),　3)　phosphorylation　of　Ser(203)　markedly　increased　in　vitro　when　recombinant　PKD1　was　incubated　with　either　PAK1　or　PAK2　in　the　presence　of　ATP.　PAK　inhibitors　did　not　interfere　with　G　protein-coupled　receptor　activation-induced　rapid　translocation　of　PKD1　to　the　plasma　membrane　but　strikingly　prevented　the　dissociation　of　PKD1　from　the　plasma　membrane　and　blunted　the　phosphorylation　of　nuclear　targets,　including　class　IIa　histone　deacetylases.　We　conclude　that　PAK-mediated　phosphorylation　of　PKD1　at　Ser(203)　triggers　its　membrane　dissociation　and　subsequent　entry　into　the　nucleus,　thereby　regulating　the　phosphorylation　of　PKD1　nuclear　targets,　including　class　IIa　histone　deacetylases.