Protein kinase A regulates cholinergic gene expression in PC12 cells: REST4 silences the silencing activity of neuron-restrictive silencer factor/REST.
The role of protein kinase A in regulating transcription of the cholinergic gene locus, which contains both the vesicular acetylcholine transporter gene and the choline acetyltransferase gene, was investigated in PC12 cells and a protein kinase A-deficient PC12 mutant, A126.1B2, in which transcription of the gene is reduced. The site of action of protein kinase A was localized to a neuron-restrictive silencer element/repressor element 1 (NRSE/RE-1) sequence within the cholinergic gene. Neuron-restrictive silencer factor (NRSF)/RE-1-silencing transcription factor (REST), the transcription factor which binds to NRSE/RE-1, was expressed at similar levels in both PC12 and A126.1B2 cells. Although nuclear extracts containing NRSF/REST from A126.1B2 exhibited binding to NRSE/RE-1, nuclear extracts from PC12 cells did not. The NRSF/REST isoform REST4 was expressed in PC12 cells but not in A126.1B2. REST4 inhibited binding of NRSF/REST to NRSE/RE-1 as determined by gel mobility shift assays. Coimmunoprecipitation was used to demonstrate interaction between NRSF/REST and REST4. Expression of recombinant REST4 in A126.1B2 was sufficient to transcriptionally activate the cholinergic gene locus. Thus, in PC12 cells, protein kinase A promotes the production of REST4, which inhibits repression of the cholinergic gene locus by NRSF/REST.
Shimojo
M
M
Department of Biochemistry, University of Kentucky, Lexington 40536-0298, USA.
Paquette
A J
AJ
Anderson
D J
DJ
Hersh
L B
LB
D000110:Acetylcholinesterase / Q000096:biosynthesis / Q000235:genetics*; D017398:Alternative Splicing; D000818:Animals; D002352:Carrier Proteins / Q000096:biosynthesis / Q000235:genetics*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D020868:Gene Silencing; D017930:Genes, Reporter; D026901:Membrane Transport Proteins; D009474:Neurons / Q000502:physiology*; D016716:PC12 Cells; D011485:Protein Binding; D051381:Rats; D012097:Repressor Proteins / Q000235:genetics* / Q000378:metabolism; D014157:Transcription Factors; D050494:Vesicular Acetylcholine Transport Proteins; D033921:Vesicular Transport Proteins; D000099229:RE1-Silencing Transcription Factor
D000110:Acetylcholinesterase; D002352:Carrier Proteins; D017868:Cyclic AMP-Dependent Protein Kinases; D026901:Membrane Transport Proteins; D012097:Repressor Proteins; D014157:Transcription Factors; D050494:Vesicular Acetylcholine Transport Proteins; D033921:Vesicular Transport Proteins; D000099229:RE1-Silencing Transcription Factor; C493461:Slc18a3 protein, rat