R-Ras controls axon specification upstream of glycogen synthase kinase-3beta through integrin-linked kinase.
The initial event in establishing a polarized neuron is the specification of a single axon. Spatially regulated glycogen synthase kinase-3beta (GSK-3beta) activity is critical for specifying axon-dendrite fate; however, the upstream signaling of GSK-3beta in the determination of neuronal polarity still remains obscure. Here, we found that, in cultured hippocampal neurons, the small GTPase R-Ras selectively localized in a single neurite of stage 2 neurons and that its activity increased after plating and peaked between stages 2 and 3. Ectopic expression of R-Ras induced global inactivation of GSK-3beta and formation of multiple axons, whereas knockdown of endogenous R-Ras by RNA interference blocked GSK-3beta inactivation and axon formation. GSK-3beta inactivation and axon formation by R-Ras required integrin-linked kinase (ILK), and subcellular localization of ILK was strictly regulated by R-Ras-mediated phosphatidylinositol 3-kinase activity. In addition, membrane targeting of ILK was sufficient to inactivate GSK-3beta and to form multiple axons. Our study demonstrates a novel role of R-Ras and ILK upstream of GSK-3beta in the regulation of neuronal polarity.
Oinuma
Izumi
I
Laboratory of Molecular Neurobiology, Graduate School of Biostudies, Kyoto University, Sakyo-ku, Kyoto 606-8502, Japan.
Katoh
Hironori
H
Negishi
Manabu
M
D000818:Animals; D001369:Axons / Q000378:metabolism*; D019556:COS Cells; D002460:Cell Line; D002522:Chlorocebus aethiops; D003712:Dendrites / Q000378:metabolism; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism*; D000071679:Glycogen Synthase Kinase 3 beta; D006624:Hippocampus / Q000378:metabolism; D006801:Humans; D009474:Neurons / Q000378:metabolism; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D034622:RNA Interference; D013997:Time Factors; D018631:ras Proteins / Q000378:metabolism*; D000099200:Scaffold Protein ILK
D038362:Glycogen Synthase Kinase 3; D000071679:Glycogen Synthase Kinase 3 beta; D017346:Protein Serine-Threonine Kinases; D018631:ras Proteins; D000099200:Scaffold Protein ILK; C000605506:GSK3B protein, human