From transcription profile to expression: the signaling repertoire of the SK-N-MC neuroepithelioma cell-line.
SK-N-MC neuroepithelioma cells are routinely cultured and widely used as a model system in biochemical and pharmacological experiments. To clarify the gene expression patterns of SK-N-MC cells with respect to G protein-coupled receptors and signaling network components, we describe in this report the transcription profile of the cell line. Following the traditional pathway from genome to proteome, selected examples are further examined at the level of protein expression and by functional assays. cRNA targets derived from total RNA extracts were hybridized to Affymetrix Human Genome U133A GeneChip arrays, and the data were analyzed and grouped according to functional aspects. Results obtained for neuropeptide Y (NPY) Y1, Y5, and orexin Ox1 receptors were confirmed by RT-PCR. It is surprising that we found the presence of both NPY receptor subtypes and the absence of the orexin receptor at the mRNA level. Receptor-binding experiments confirmed NPY binding of the Y1 receptor in the nanomolar range but gave no evidence for high expression levels of Y5 receptor subtypes on the cell surface. Protein expression was assayed with immunoblots by using antibodies directed against selected Galpha protein subunits. The presence of at least Galphas, Galphai3, and Galphai2 subunits was indicated.
Bader
Jürgen E
JE
Faculty of Biosciences, Pharmacy and Psychology, Institute of Biochemistry, University of Leipzig, Leipzig, Germany.
Deckert
Cornelia M
CM
Koglin
Norman
N
Pluder
Franka
F
Mörl
Karin
K
Koczan
Dirk
D
Thiesen
Hans-Jürgen
HJ
Beck-Sickinger
Annette G
AG
D001665:Binding Sites; D015153:Blotting, Western; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D015151:Immunoblotting; D020128:Inhibitory Concentration 50; D018241:Neuroectodermal Tumors, Primitive, Peripheral / Q000378:metabolism*; D020411:Oligonucleotide Array Sequence Analysis; D064446:Orexin Receptors; D010455:Peptides / Q000737:chemistry; D011485:Protein Binding; D012313:RNA / Q000737:chemistry; D018075:RNA, Complementary / Q000378:metabolism; D043562:Receptors, G-Protein-Coupled; D018013:Receptors, Neuropeptide / Q000378:metabolism; D017476:Receptors, Neuropeptide Y / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction; D014158:Transcription, Genetic
D064446:Orexin Receptors; D010455:Peptides; D018075:RNA, Complementary; D043562:Receptors, G-Protein-Coupled; D018013:Receptors, Neuropeptide; D017476:Receptors, Neuropeptide Y; C097212:neuropeptide Y-Y1 receptor; C100568:neuropeptide Y5 receptor; D012313:RNA