Neuritin (cpg15) enhances the differentiating effect of NGF on neuronal PC12 cells.
Neuritin is a small, highly conserved GPI-anchored protein involved in neurite outgrowth. We have analyzed the involvement of neuritin in NGF-induced differentiation of PC12 cells by investigating the time-course of neuritin expression, the effects of its overexpression or silencing, and the possible mechanisms of its regulation and action. Real-time PCR analysis has shown that neuritin gene is upregulated by NGF in PC12 cells hours before neurite outgrowth becomes appreciable. PC12 cells transfected with a plasmid expressing neuritin display a significant increase in the response to NGF: 1) in the levels of SMI312 positive phosphorylated neurofilament proteins (markers for axonal processes) and tyrosine hydroxylase; 2) in the percentage of cells bearing neurites; as well as 3) in the average length of neurites when compared to control cells. On the contrary, neuritin silencing significantly reduces neurite outgrowth. These data suggest that neuritin is a modulator of NGF-induced neurite extension in PC12 cells. We also showed that neuritin potentiated the NGF-induced differentiation of PC12 cells without affecting TrkA or EGF receptor mRNAs expression. Moreover, the S-methylisothiourea (MIU), a potent inhibitor of inducible nitric oxide synthases, partially counteracts the NGF-mediated neuritin induction. These data suggest that NGF regulates neuritin expression in PC12 cells via the signaling pathway triggered by NO. This study reports the first evidence that neuritin plays a role in modulating neurite outgrowth during the progression of NGF-induced differentiation of PC12 cells. PC12 cells could be considered a valuable model to unravel the mechanism of action of neuritin on neurite outgrowth. (c) 2007 Wiley-Liss, Inc.
Cappelletti
Graziella
G
Department of Biology, University of Milan, Milan, Italy.
Galbiati
Mariarita
M
Ronchi
Cristina
C
Maggioni
Maria Grazia
MG
Onesto
Elisa
E
Poletti
Angelo
A
D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects*; D004347:Drug Interactions; D004791:Enzyme Inhibitors / Q000494:pharmacology; D058851:GPI-Linked Proteins; D005786:Gene Expression Regulation / Q000187:drug effects*; D049452:Green Fluorescent Proteins / Q000378:metabolism; D007550:Isothiuronium / Q000031:analogs & derivatives / Q000494:pharmacology; D008565:Membrane Proteins / Q000502:physiology*; D020932:Nerve Growth Factor / Q000494:pharmacology*; D009419:Nerve Tissue Proteins / Q000502:physiology*; D016900:Neurofilament Proteins / Q000378:metabolism; D009474:Neurons / Q000502:physiology*; D016716:PC12 Cells / Q000187:drug effects; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D013997:Time Factors; D014162:Transfection / Q000379:methods; D014446:Tyrosine 3-Monooxygenase / Q000378:metabolism
D004791:Enzyme Inhibitors; D058851:GPI-Linked Proteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D016900:Neurofilament Proteins; C484377:Nrn1 protein, mouse; D012333:RNA, Messenger; C061195:neurofilament protein M; D049452:Green Fluorescent Proteins; D007550:Isothiuronium; D020932:Nerve Growth Factor; D014446:Tyrosine 3-Monooxygenase; C027744:S-methylisothiopseudouronium