title: Sphingosine kinase 1/S1P pathway involvement in the GDNF-induced GAP43 transcription.
abstract: Glial cell line-derived neurotrophic factor (GDNF) is important for the development and maintenance of dopamine neurons (Lin et al. [1993] Science 260: 1130-1132). GDNF is neuroprotective in animal models of Parkinson disease, where dopamine neurons show selective degeneration. We previously reported GDNF-induced SPHK1 gene expression in a neuroblastoma cell line, TGW (Murakami et al. [2007] J Neurochem 102: 1585-1594). In the present study, we focused on the regulatory mechanism of GAP43 (GDNF-induced neuronal phenotype) transcription to further elucidate physiological roles of GDNF-induced SPHK1 expression and activity. Stable wild-type (SPHK1-WT) but not dominant-negative SPHK1 (SPHK1-DN) overexpression increased both control- and GDNF-induced GAP43 expression. SPHK1-WT cells showed enhanced GDNF-induced sphingosine 1-phosphate (S1P) secretion compared with mock- and SPHK1-DN cells. Exogenous S1P also increased GAP43 expression. In TGW cells, PD98059, a MEK inhibitor, but not SB203580 (a p38 MAPK inhibitor) and LY294002 (a PI3K inhibitor) inhibited GDNF-induced GAP43 expression, suggesting the MEK/ERK pathway has a major role in GDNF-induced GAP43 transcription. A G-protein-coupled receptor inhibitor, pertussis toxin, and S1P(1) and S1P(3) receptor antagonists (VPC23019 and CAY10444) also inhibited ERK activation. Moreover, both S1P1 and S1P3 were serine-phosphorylated by GDNF, suggesting their activated states. C/EBPβ transcription factor was induced by GDNF, and DNA pull-down and chromatin immunoprecipitation assays revealed the C/EBP binding site between -131 bp and -98  bp from the first exon of GAP43. Taken together, our results showed that in TGW cells, GDNF increased SPHK1 transcription, leading to the production and secretion of S1P. Through MEK/ERK pathway, S1P stimulates GAP43 transcription with increased binding of C/EBPβ to the 5'-promoter.
authors.affiliation: Department of Oral Disease Research, National Center for Geriatrics and Gerontology, Obu, Japan.
authors.forename: Masashi
authors.initials: M
authors.lastname: Murakami
authors.forename: Hiromi
authors.initials: H
authors.lastname: Ito
authors.forename: Kazumi
authors.initials: K
authors.lastname: Hagiwara
authors.forename: Misa
authors.initials: M
authors.lastname: Kobayashi
authors.forename: Asuka
authors.initials: A
authors.lastname: Hoshikawa
authors.forename: Akira
authors.initials: A
authors.lastname: Takagi
authors.forename: Tetsuhito
authors.initials: T
authors.lastname: Kojima
authors.forename: Keiko
authors.initials: K
authors.lastname: Tamiya-Koizumi
authors.forename: Sayaka
authors.initials: S
authors.lastname: Sobue
authors.forename: Masatoshi
authors.initials: M
authors.lastname: Ichihara
authors.forename: Motoshi
authors.initials: M
authors.lastname: Suzuki
authors.forename: Yoshiko
authors.initials: Y
authors.lastname: Banno
authors.forename: Yoshinori
authors.initials: Y
authors.lastname: Nozawa
authors.forename: Takashi
authors.initials: T
authors.lastname: Murate
mesh_terms: D001483:Base Sequence; D015153:Blotting, Western; D045744:Cell Line, Tumor; D047369:Chromatin Immunoprecipitation; D003001:Cloning, Molecular; D017931:DNA Primers; D019922:GAP-43 Protein / Q000235:genetics*; D051100:Glial Cell Line-Derived Neurotrophic Factor / Q000378:metabolism*; D006801:Humans; D008246:Lysophospholipids / Q000378:metabolism*; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D015398:Signal Transduction; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000378:metabolism; D014158:Transcription, Genetic; D000099157:Sphingosine Kinase
chemical_list: D017931:DNA Primers; D019922:GAP-43 Protein; D051100:Glial Cell Line-Derived Neurotrophic Factor; D008246:Lysophospholipids; D017853:Phosphotransferases (Alcohol Group Acceptor); D013110:Sphingosine; D000099157:Sphingosine Kinase; C060506:sphingosine 1-phosphate