An AP-2 binding sequence within exon 1 of human and porcine choline acetyltransferase genes enhances transcription in neural cells.
The gene for choline acetyltransferase, synthesizing acetylcholine, is induced by several neurotrophic factors. A role for AP-2 in enhancing this transcription and limiting it to neural cells is strongly suggested. Previous studies demonstrated that base pairs +465-727 within the untranslated exon 1 of the porcine gene enhanced the expression of a reporter gene transfected into PC-12 cells. Deletion and mutation experiments indicate that base pairs +465-472 (CCGCGGGG) in the porcine gene, or +307-314 (CCTCGGGG) in the human sequence, were necessary and sufficient for increased gene expression in cholinergic or adrenergic but not liver cells. Constructs containing active sequences, but not inactive mutated sequences, specifically bind nuclear proteins from neuroblastoma cells, but not liver cells, in gel shift experiments. The human and porcine sequences are in agreement with an AP-2 consensus binding sequence, a nuclear transcription factor expressed only in cells derived from the neural crest. Gel shift experiments using recombinant AP-2 confirm this identification. AP-2 antibody further retarded the mobility of these DNA-nuclear extract or DNA-AP-2 complexes. These results support the importance of this AP-2 binding sequence in enhancing and limiting choline acetyltransferase expression in neural cells.
Baskin
F
F
Department of Neurology, The University of Texas Southwestern Medical Center at Dallas, 75235, U.S.A.
Li
Y P
YP
Hersh
L B
LB
Davis
R M
RM
Rosenberg
R N
RN
D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D002795:Choline O-Acetyltransferase / Q000235:genetics*; D005091:Exons / Q000502:physiology*; D006801:Humans; D008969:Molecular Sequence Data; D009474:Neurons / Q000201:enzymology*; D009687:Nuclear Proteins / Q000378:metabolism; D009841:Oligonucleotides / Q000138:chemical synthesis / Q000378:metabolism; D010957:Plasmids; D011485:Protein Binding; D051381:Rats; D013552:Swine; D018808:Transcription Factor AP-1 / Q000378:metabolism*; D014158:Transcription, Genetic / Q000502:physiology*; D014162:Transfection
D009687:Nuclear Proteins; D009841:Oligonucleotides; D018808:Transcription Factor AP-1; D002795:Choline O-Acetyltransferase