The requirement of Ras and Rap1 for the activation of ERKs by cAMP, PACAP, and KCl in cerebellar granule cells.
In cerebellar granule cells, the mitogen-activated protein kinase (MAPK) or extracellular signal-regulated kinase (ERK) cascade mediates multiple functions, including proliferation, differentiation, and survival. In these cells, ERKs are activated by diverse stimuli, including cyclic adenosine monophosphate (cAMP), pituitary adenylate cyclase activating protein (PACAP), depolarization induced by elevated extracellular potassium (KCl), and the neurotrophin brain-derived neurotrophic factor. Extensive studies in neuronal cell lines have implicated the small G proteins Ras and Rap1 in the activation of ERKs by cAMP, PACAP, and KCl. However, the requirement of Ras and Rap1 in these pathways in cerebellar granule cells has not been addressed. In this study, we utilize multiple biochemical assays to determine the mechanisms of action and requirement of Ras and Rap1 in cultured cerebellar granule cells. We show that both Ras and Rap1 can be activated by cAMP or PACAP via protein kinase (PKA)-dependent mechanisms. KCl activation of Ras also required PKA. Using both adenoviral and transgenic approaches, we show that Ras plays a major role in ERK activation by cAMP, PACAP, and KCl, while Rap1 also mediates activation of a selective membrane-associated pool of ERKs. Furthermore, Rap1, but not Ras, activation by PKA appears to require the action of Src family kinases.
Obara
Yutaro
Y
Vollum Institute, L474, Oregon Health and Science University, Portland, Oregon 97239, USA.
Horgan
Angela M
AM
Stork
Philip J S
PJ
D000818:Animals; D002478:Cells, Cultured; D002531:Cerebellum / Q000166:cytology / Q000201:enzymology; D000242:Cyclic AMP / Q000378:metabolism* / Q000494:pharmacology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000187:drug effects / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism*; D051379:Mice; D008822:Mice, Transgenic; D009474:Neurons / Q000201:enzymology*; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide / Q000378:metabolism* / Q000494:pharmacology; D011189:Potassium Chloride / Q000494:pharmacology; D051381:Rats; D020827:rap1 GTP-Binding Proteins / Q000378:metabolism*; D018631:ras Proteins / Q000378:metabolism*
D051219:Pituitary Adenylate Cyclase-Activating Polypeptide; D011189:Potassium Chloride; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D048049:Extracellular Signal-Regulated MAP Kinases; D020827:rap1 GTP-Binding Proteins; D018631:ras Proteins