title: Nerve growth factor down-regulates the expression of norepinephrine transporter in rat pheochromocytoma (PC12) cells.
abstract: Functional expression of norepinephrine transporter (NET) and its regulation were examined in rat pheochromocytoma cell line, PC12. Nerve growth factor (NGF) decreased [3H]-norepinephrine (NE) uptake in association with a decrease in NET mRNA levels. On the other hand, levels of tyrosine hydroxylase mRNA increased in PC12 cells treated with NGF for 4-24 h, while Oct-2 mRNA levels decreased at 4 h with NGF then recovered for 8-24 h in the presence of NGF. Both bFGF and EGF reduced [3H]NE uptake, although they failed to affect NET mRNA levels. To examine the NET transcriptional regulation, we identified the 5'-noncoding region of rat NET mRNA by the rapid amplification of cDNA end (RACE) method and cloned the 5'-flanking region of NET gene. The newly identified exon encodes the untranslated region of rat NET mRNA upstream of the known 5'-region including ATG start codon. Constructs having green fluorescent protein (GFP) as reporter were made with the cloned NET gene, and promoter activity was examined in CHO and SK-N-SH cells transiently transfected and in PC12 cells stably transfected with NET-GFP constructs. The results indicate that the 2.1 kb NET flanking region displays promoter activity and is responsible for the NGF-induced down-regulation of NET expression.
authors.affiliation: Department of Pharmacology, Hiroshima University Faculty of Dentistry, Kasumi 1-2-3, Minamiku, 734-8553, Hiroshima, Japan.
authors.forename: T
authors.initials: T
authors.lastname: Ikeda
authors.forename: S
authors.initials: S
authors.lastname: Kitayama
authors.forename: K
authors.initials: K
authors.lastname: Morita
authors.forename: T
authors.initials: T
authors.lastname: Dohi
mesh_terms: D020121:5' Untranslated Regions / Q000235:genetics; D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D002352:Carrier Proteins / Q000235:genetics*; D003001:Cloning, Molecular; D006224:Cricetinae; D004268:DNA-Binding Proteins / Q000235:genetics; D015536:Down-Regulation / Q000235:genetics; D015870:Gene Expression / Q000187:drug effects; D008969:Molecular Sequence Data; D020932:Nerve Growth Factor / Q000494:pharmacology*; D009474:Neurons / Q000502:physiology*; D009638:Norepinephrine / Q000493:pharmacokinetics; D050484:Norepinephrine Plasma Membrane Transport Proteins; D050813:Octamer Transcription Factor-2; D016716:PC12 Cells; D012333:RNA, Messenger / Q000032:analysis; D011869:Radioligand Assay; D051381:Rats; D013566:Sympathomimetics / Q000493:pharmacokinetics; D027981:Symporters; D014157:Transcription Factors / Q000235:genetics; D015533:Transcriptional Activation / Q000502:physiology; D014316:Tritium; D014446:Tyrosine 3-Monooxygenase / Q000235:genetics
chemical_list: D020121:5' Untranslated Regions; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D050484:Norepinephrine Plasma Membrane Transport Proteins; D050813:Octamer Transcription Factor-2; C489205:Pou2f2 protein, rat; D012333:RNA, Messenger; C493226:Slc6a2 protein, rat; D013566:Sympathomimetics; D027981:Symporters; D014157:Transcription Factors; D014316:Tritium; D020932:Nerve Growth Factor; D014446:Tyrosine 3-Monooxygenase; D009638:Norepinephrine