title: The Neuro-2a neuroblastoma cell line expresses [Met]-enkephalin and vasopressin mRNA and peptide.
abstract: Mouse neuroblastoma Neuro-2a cells were examined for the expression of pro-enkephalin mRNA, protein, and Met-enkephalin ([Met]-Enk) peptide. Reverse transcriptase/polymerase chain reaction (RT/PCR) and in situ hybridization demonstrated the presence of pro-enkephalin mRNA in these cells. Immunocytochemistry using an antibody which recognizes pro-enkephalin and high pressure liquid chromatography (HPLC) followed by radioimmunoassay indicated that pro-enkephalin was synthesized in these cells and processed to yield the bioactive pentapeptide, [Met]-Enk. Furthermore, release studies showed that the [Met]-Enk was secreted from these cells with high K+ stimulation. Using double labeling, in situ hybridization combined with immunocytochemistry, we demonstrated that prohormone convertase 2 (PC2) mRNA is colocalized with pro-enkephalin in the same Neuro-2a cells, suggesting that this enzyme may be responsible for processing this precursor. we also showed the presence of vasopressin mRNA and arginine-vasopressin peptide in these cells using in situ hybridization and immunocytochemistry, respectively. Thus, the Neuro-2a cells are a multiple neuropeptide-producing cell line and an excellent model for studying the mechanisms involved in the synthesis, intracellular targeting and processing of endogenous pro-enkephalin and pro-vasopressin, as well as other transfected neuropeptide precursors.
authors.affiliation: National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: A M
authors.initials: AM
authors.lastname: Bamberger
authors.forename: L P
authors.initials: LP
authors.lastname: Pu
authors.forename: D R
authors.initials: DR
authors.lastname: Cool
authors.forename: Y P
authors.initials: YP
authors.lastname: Loh
mesh_terms: D000818:Animals; D001127:Arginine Vasopressin / Q000032:analysis; D002851:Chromatography, High Pressure Liquid; D004744:Enkephalin, Methionine / Q000235:genetics* / Q000378:metabolism; D004745:Enkephalins / Q000032:analysis / Q000235:genetics; D015870:Gene Expression; D007150:Immunohistochemistry; D017403:In Situ Hybridization; D051379:Mice; D009447:Neuroblastoma / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D045706:Proprotein Convertase 2; D011498:Protein Precursors / Q000032:analysis / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis / Q000378:metabolism*; D012194:RNA-Directed DNA Polymerase; D013381:Subtilisins / Q000235:genetics; D014407:Tumor Cells, Cultured; D014667:Vasopressins / Q000032:analysis / Q000235:genetics*
chemical_list: D004745:Enkephalins; D011498:Protein Precursors; D012333:RNA, Messenger; C029992:proenkephalin; D014667:Vasopressins; D001127:Arginine Vasopressin; D004744:Enkephalin, Methionine; D012194:RNA-Directed DNA Polymerase; D013381:Subtilisins; D045706:Proprotein Convertase 2