Coordinated up-regulation of choline acetyltransferase and vesicular acetylcholine transporter gene expression by the retinoic acid receptor alpha, cAMP, and leukemia inhibitory factor/ciliary neurotrophic factor signaling pathways in a murine septal cell line

Abstract

The proteins responsible for acetylcholine (ACh) synthesis (choline acetyltransferase, ChAT) and storage (vesicular ACh transporter, VAChT) are encoded by two closely linked genes in vertebrates, with the VAChT coding sequence contained within the first intron of the ChAT gene. This unusual genomic organization suggests that the transcription of these two genes is coordinately regulated. Using Northern analysis we studied the modulation of ChAT and VAChT expression in a murine septal cell line (SN56) by three groups of agents: retinoids, trophic factors belonging to the leukemia inhibitory factor/ciliary neurotrophic factor (LIF/CNTF) family, and cAMP. All-trans-retinoic acid increased both ChAT and VAChT mRNA levels in SN56 cells up to 3.5-fold, and elevated intracellular ACh levels by 2.5-fold. This effect was mimicked by a retinoic acid receptor alpha (RAR alpha) agonist (Ro 40-6055) and prevented by a specific antagonist (Ro 41-5253), indicating that it was mediated by RAR alpha. ChAT- and VAChT-specific transcripts were also induced (up to 3-fold) by treatment with CNTF or LIF (20 ng/ml, 48 h), as well as by dibutyryl cAMP (1 mM). All these agents increased the ACh level in the cells (up to 2.5-fold). Dibutyryl cAMP had a greater effect on the level of VAChT mRNA (4-fold induction) than on the level of ChAT mRNA (2-fold induction), suggesting a quantitatively differential transcriptional regulation of the two genes by the cAMP pathway. The effects of the three groups of agents studied on ChAT and VAChT mRNA levels were additive, pointing to several independent mechanisms by which the cholinergic properties of septal neurons can be modulated.