delta Ca(2+)/Calmodulin-dependent protein kinase II isozyme-specific induction of neurite outgrowth in P19 embryonal carcinoma cells.
Ca(2+)/calmodulin-dependent protein kinase II (CaMK-II) has been linked to the induction of differentiation in preneuronal cells. In these cells, delta isozymes represent the majority of CaMK-IIs expressed and are activated by differentiation stimuli. To determine whether delta CaMK-IIs are causative or coincident with in vitro differentiation, we overexpressed wild-type, constitutively active, and C-terminal domains of delta and gamma CaMK-II isozymes in mouse P19 and NIH/3T3 cells using high-efficiency transfections. At 1-2 days after transfection, only constitutively active delta CaMK-II isozymes induced branched cellular extensions in both cell types. In P19 cells, retinoic acid induced neurite extensions after 3-4 days; these extensions were coincident with a fourfold increase in endogenous CaMK-II activity. Extensions induced by both retinoic acid and delta CaMK-IIs contained class III beta-tubulin in a discontinuous or beaded pattern. C-terminal CaMK-II constructs disrupted the ability of endogenous CaMK-II to autophosphorylate and blocked retinoic acid-induced differentiation. delta CaMK-II was found along extensions, whereas gamma CaMK-II exhibited a more diffuse, cytosolic localization. These data not only support an extranuclear role for CaMK-II in promoting neurite outgrowth, but also demonstrate CaMK-II isozyme specificity in these early steps of neuronal differentiation.
Johnson
L D
LD
Department of Biology and Massey Cancer Center, Virginia Commonwealth University, Richmond, Virginia 23284-2012, USA.
Willoughby
C A
CA
Burke
S H
SH
Paik
D S
DS
Jenkins
K J
KJ
Tombes
R M
RM
D016475:3T3 Cells; D000818:Animals; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D002454:Cell Differentiation / Q000187:drug effects / Q000235:genetics; D002455:Cell Division / Q000187:drug effects / Q000235:genetics; D054278:Embryonal Carcinoma Stem Cells; D004789:Enzyme Activation / Q000187:drug effects; D015870:Gene Expression; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D051379:Mice; D014411:Neoplastic Stem Cells / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D016501:Neurites / Q000187:drug effects / Q000201:enzymology*; D009474:Neurons / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D014162:Transfection; D014212:Tretinoin / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D007527:Isoenzymes; D014212:Tretinoin; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases