Cdk5/p35 and Rho-kinase mediate ephrin-A5-induced signaling in retinal ganglion cells.
Ephrin-As are repulsive axonal guidance cues that regulate retinotectal projection. EphA tyrosine kinases, which are the receptors of ephrin-As, activate signaling cascades leading to cytosckeleton reorganization. Here, we address the role of cyclin-dependent kinase (Cdk) 5 in Eph receptor signaling induced by ephrin-A5. Ephrin-A5 induced a cell morphological response in PC-3M cells that endogenously express Cdk5 and EphA2, a receptor for ephrin-A5. This response was augmented by the transfection of p35, which is a neuronal regulator of Cdk5. While the morphological response of native PC-3M cells was not affected by olomoucine, an inhibitor of Cdk, the response was inhibited in the p35-transfected cells. In retinal ganglion cells, either olomoucine at 20 microM or Y-27632 at 10 microM, an inhibitor of Rho-kinase/ROKalpha/ROCKII, showed maximum inhibitory effect against ephrin-A5 (10 microg/ml)-induced growth cone collapse. Combined application of olomoucine and Y-27632 further suppressed the ephrin-A5-induced response. Ephrin-A5 evoked phosphorylation of Cdk5 at Tyr15 and tau, a substrate of Cdk5 in retinal growth cones. Recombinant herpes simplex virus expressing Cdk5 mutant (kinase-negative or Tyr15 to Ala) showed a dominant-negative effect on the ephrin-A5-induced growth cone collapse. These findings demonstrate that both Cdk5 and the Rho kinase pathway independently contribute to the downstream of ephrin-A-induced signaling in retinal ganglion cells.
Cheng
Qi
Q
Department of Molecular Pharmacology and Neurobiology, Yokohama City University School of Medicine, Yokohama, 236-0004, Japan.
Sasaki
Yukio
Y
Shoji
Masayuki
M
Sugiyama
Yoshinobu
Y
Tanaka
Hideaki
H
Nakayama
Takashi
T
Mizuki
Nobuhisa
N
Nakamura
Fumio
F
Takei
Kohtaro
K
Goshima
Yoshio
Y
D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D002460:Cell Line; D048429:Cell Size / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D002642:Chick Embryo; D051360:Cyclin-Dependent Kinase 5; D018844:Cyclin-Dependent Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D036386:Ephrin-A5 / Q000378:metabolism* / Q000494:pharmacology; D020439:Growth Cones / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D009154:Mutation / Q000235:genetics; D009419:Nerve Tissue Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D036104:Receptor, EphA2 / Q000187:drug effects / Q000378:metabolism; D012160:Retina / Q000166:cytology / Q000196:embryology / Q000378:metabolism; D012165:Retinal Ganglion Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D013477:Superior Colliculi / Q000166:cytology / Q000196:embryology / Q000378:metabolism; D014795:Visual Pathways / Q000166:cytology / Q000196:embryology / Q000378:metabolism; D054460:rho-Associated Kinases
D004791:Enzyme Inhibitors; D036386:Ephrin-A5; D047908:Intracellular Signaling Peptides and Proteins; D009419:Nerve Tissue Proteins; C089495:neuronal Cdk5 activator (p25-p35); D036104:Receptor, EphA2; D051360:Cyclin-Dependent Kinase 5; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; C495905:CDK5 protein, human; D018844:Cyclin-Dependent Kinases