Synapse-specific and neuregulin-induced transcription require an ets site that binds GABPalpha/GABPbeta

Localization of acetylcholine receptors (AChRs) to neuromuscular synapses is mediated by multiple pathways. Agrin, which is the signal for one pathway, stimulates a redistribution of previously unlocalized AChRs to synaptic sites. The signal for a second pathway is not known, but this signal stimulates selective transcription of AChR genes in myofiber nuclei located near the synaptic site. Neuregulin (NRG) is a good candidate for the extracellular signal that induces synapse-specific gene expression, since NRG is concentrated at synaptic sites and activates AChR gene expression in cultured muscle cells. Previous studies have demonstrated that 181 bp of 5' flanking DNA from the AChR delta-subunit gene are sufficient to confer synapse-specific transcription in transgenic mice and NRG responsiveness in cultured muscle cells, but the critical sequences within this cis-acting regulatory region have not been identified. We transfected AChR delta-subunit-hGH gene fusions into a muscle cell line, and we show that a potential binding site for Ets proteins is required for NRG-induced gene expression. Furthermore, we produced transgenic mice carrying AChR delta-subunit-hGH gene fusions with a mutation in this NRG-response element (NRE), and we show that this NRE is necessary for synapse-specific transcription in mice. The NRE binds proteins in myotube nuclear extracts, and nucleotides that are important for NRG responsiveness are likewise critical for formation of the protein-DNA complex. This complex contains GABPalpha, an Ets protein, and GABPbeta, a protein that lacks an Ets domain but dimerizes with GABPalpha, because formation of the protein-DNA complex is inhibited by antibodies to either GABPalpha or GABPbeta. These results demonstrate that synapse-specific and NRG-induced gene expression require an Ets-binding site and suggest that GABPalpha/GABPbeta mediates the transcriptional response of the AChR delta-subunit gene to synaptic signals, including NRG.