The Section on Molecular Neuroscience studies the cis-acting elements and trans-acting factors responsible for cell-specific neurotransmitter expression, or 'chemical coding' in the central nervous system (CNS). The SMN focuses on genes encoding proteins responsible for cholinergic and monoaminergic phenotypes, and vasoactive intestinal polypeptide (VIP), a developmentally important neuropeptide co-expressed with either acetylcholine or monoamines in neuroendocrine cells. The ultimate goal of the chemical coding project is molecular understanding of nervous system development, and use of neurotransmitter-specific gene function to restore neurotransmitter balance in animal models of human CNS disease. 1. We previously showed that the mammalian cholinergic gene locus (CGL) encodes choline acetyltransferase (ChAT) and the vesicular acetylcholine transporter (VAChT) as physically contiguous genes. We have now discovered that VAChT and ChAT are transcribed at separate start sites, and have identified cis-acting elements potentially regulating both transcripts, including a non-neuronal silencer element (NRSE). A human CGL 'mini-locus' was constructed and expressed in transgenic mice. Additional CGL mini-gene constructs will be employed to identify elements responsible for CNS vs. peripheral CGL expression, and for neurotrophin- regulated CGL and VIP gene transcription (see below). 2. The VIP gene contains three discrete but functionally linked domains responsible for cell-specific VIP expression. One is an Oct-1-like consensus sequence (the VIP TSE) that we hypothesize binds a regulatory protein expressed only in VIPergic neurons. We plan to identify the neuron-specific factor that binds to the TSE and interacts with downstream regulatory factors to allow correct expression of the VIP gene. We have identified PACAP as a neuropeptide neurotrophic factor that robustly regulates VIP expression through a calcineurin- dependent pathway in neuroendocrine cells, and will explore its relevance to cell-specific VIP gene regulation in vivo. 3. The vesicular amine transporters VAChT and VMAT are co-expressed, with later reciprocal extinction, in sympathetic and parasympathetic neurons respectively, while in the CNS they are mutually exclusively expressed from early development. Perinatally-specific VMAT2 expression in sensory relay neurons of the brain has been documented, and may be important in influencing perinatal sensory cortical development. Analysis of the VMAT1 and VMAT2 promoters along with those for VIP and the CGL, will define cis-regulatory elements mediating differential neuronal expression of these genes, as a model for the regulatory cascade responsible for full chemical coding in the mature nervous system.
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