Src-mediated activation of the human neurotensin/neuromedin N promoter.
BACKGROUND
Expression of the gene encoding the neurotensin/neuromedin N (NT/N) is developmentally regulated in the gut in a distinctive temporal and spatial fashion. Src kinase, a nonreceptor tyrosine kinase, has been implicated in the growth and differentiation of various tissues; its role in gut differentiation is not known. The purpose of this study was to determine whether the Src signaling pathway plays a role in the activation of the human NT/N promoter.


METHODS
Caco-2 cells, a human colon cancer cell line that can differentiate to a small bowel phenotype, were transiently transfected with human NT/N promoter fragments linked to luciferase and various amounts of Src expression plasmids or dominant negative Raf; luciferase and beta-galactosidase activities were measured after 48 hours.


RESULTS
Cotransfection of Src resulted in an approximate eightfold increase of NT/N promoter activity; mutation of a proximal activating protein-1/cyclic adenosine monophosphate responsive element site resulted in a dramatic decrease of Src-mediated NT/N induction. Cotransfection with a dominant negative Raf plasmid partially blocked Src-mediated NT/N activation.


CONCLUSIONS
Src increases NT/N promoter activity in Caco-2 cells acting, in part, through a proximal AP-1/CRE promoter element. In addition, Src regulation of the NT/N promoter appears to be mediated through a Raf-dependent pathway. We propose that Src may play a role in tissue-specific gene expression in the gut.
Banker
N A
NA
Department of Surgery, University of Texas Medical Branch, Galveston 77555-0533, USA.
Hellmich
M R
MR
Kim
H J
HJ
Townsend
C M
CM
Evers
B M
BM
D000081247:CSK Tyrosine-Protein Kinase; D003110:Colonic Neoplasms; D005786:Gene Expression Regulation; D006801:Humans; D008156:Luciferases / Q000096:biosynthesis; D009496:Neurotensin / Q000096:biosynthesis* / Q000235:genetics*; D010446:Peptide Fragments / Q000096:biosynthesis* / Q000235:genetics*; D011401:Promoter Regions, Genetic; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000096:biosynthesis / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D019908:Proto-Oncogene Proteins c-raf; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D014162:Transfection; D014407:Tumor Cells, Cultured; D001616:beta-Galactosidase / Q000096:biosynthesis; D018631:ras Proteins / Q000378:metabolism; D019061:src-Family Kinases
D010446:Peptide Fragments; D011518:Proto-Oncogene Proteins; D011993:Recombinant Fusion Proteins; C042090:neuromedin N; D009496:Neurotensin; D008156:Luciferases; D011505:Protein-Tyrosine Kinases; D000081247:CSK Tyrosine-Protein Kinase; D019061:src-Family Kinases; C000631346:CSK protein, human; D017346:Protein Serine-Threonine Kinases; D019908:Proto-Oncogene Proteins c-raf; D001616:beta-Galactosidase; D018631:ras Proteins