Beta1,4-N-Acetylglucosaminyltransferase III down-regulates neurite outgrowth induced by costimulation of epidermal growth factor and integrins through the Ras/ERK signaling pathway in PC12 cells.
A rat pheochromocytoma cell line (PC12), when transfected with beta1,4-N-acetylglucosaminyltransferase III (GnT-III), which catalyzes the formation of a bisecting GlcNAc structure in N-glycans, resulted in the suppression of neurite outgrowth induced by costimulation of epidermal growth factor (EGF) and integrins. The neurite outgrowth was restored by the overexpression of a constitutively activated mitogen- or extracellular signal-regulated kinase kinase-1 (MEK-1). Consistent with this, the EGF receptor (EGFR)-mediated ERK activation was blocked in GnT-III transfectants. Conversely, the overexpression of dominant negative MEK-1 or treatment with PD98059, a specific inhibitor of MEK-1, inhibited neurite outgrowth in controls transfected with mock. Furthermore GnT-III activity is required for these inhibitions, because the overexpression of a dominant negative GnT-III mutant (D321A) failed to reduce neurite outgrowth and EGFR-mediated ERK activation. Lectin blot analysis confirmed that EGFR from wild-type GnT-III transfectants had been modified by bisecting GlcNAc in its N-glycan structures. This modification led to a significant decrease in EGF binding and EGFR autophosphorylation. Collectively, the results constitute a comprehensive body of evidence to show clearly that the overexpression of GnT-III prevents neurite outgrowth induced by costimulation of EGF and integrins through the Ras/MAPK activation pathway and indicates that GnT-III may be an important regulator for cell differentiation in neural tissues.
Gu
Jianguo
J
Department of Biochemistry, Osaka University Graduate School of Medicine, B1, 2-2 Yamadaoka Suita, Osaka 565-0871, Japan. jgu@biochem.med.osaka-u.ac.jp
Zhao
Yanyang
Y
Isaji
Tomoya
T
Shibukawa
Yukinao
Y
Ihara
Hideyuki
H
Takahashi
Motoko
M
Ikeda
Yoshitaka
Y
Miyoshi
Eiji
E
Honke
Koichi
K
Taniguchi
Naoyuki
N
D000818:Animals; D002454:Cell Differentiation; D002478:Cells, Cultured; D003094:Collagen / Q000378:metabolism; D004815:Epidermal Growth Factor / Q000378:metabolism*; D006031:Glycosylation; D016023:Integrins / Q000378:metabolism*; D048369:MAP Kinase Kinase 1; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism; D017350:N-Acetylgalactosaminyltransferases / Q000378:metabolism*; D016501:Neurites / Q000378:metabolism*; D016716:PC12 Cells; D011485:Protein Binding; D051381:Rats; D015398:Signal Transduction; D014162:Transfection
D016023:Integrins; D004815:Epidermal Growth Factor; D003094:Collagen; C480161:B4GALNT3 protein, human; D017350:N-Acetylgalactosaminyltransferases; D048369:MAP Kinase Kinase 1; C482125:MAP2K1 protein, human; D020929:Mitogen-Activated Protein Kinase Kinases