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 ge ne and the choline acetyltransferase gene, was investigated in PC12 cells a nd a protein kinase A-deficient PC12 mutant, A126.1B2, in which transcripti on of the gene is reduced. The site of action of protein kinase A was local ized 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 facto r which binds to NRSE/RE-1, was expressed at similar levels in both PC12 an d A126.1B2 cells. Although nuclear extracts containing NRSF/REST from A126. 1B2 exhibited binding to NRSE/RE-1, nuclear extracts from PC12 cells did no t. 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 inter action between NRSF/REST and REST 1. Expression of recombinant REST4 in A12 6.1B2 was sufficient to transcriptionally activate the cholinergic gene loc us. Thus, in PC12 cells, protein kinase A promotes the production of REST4, which inhibits repression of the cholinergic gene locus by NRSF/REST.