The Regulation of Ca2+-Gated and Voltage-Gated K+ Channel Genes in the Nervous System

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PI: Nigel AtkinsonAbstractDifferent types of neurons must have different patterns of electrical activity for the nervous system to function properly. For example, neurons that initiate movement often "fire" high frequency bursts of action potentials, whereas neurons that control slow, rhythmic behaviors may fire only slowly, but with very precise timing. A neuron's ability to generate specific activity patterns is determined by the types and numbers of ion channels it expresses. Nerve cells pick and choose which channels to express by transcriptional regulation. Our understanding of how cells make this decision is very immature. Furthermore, our ability to inspect the sequence of a gene and to then predict its expression pattern is nonexistent. Presently, the purpose of DNA sequences that regulate gene expression (control elements) can only be empirically ascertained. Dr. Atkinson and his students will utilize the genome project, evolutionary studies and functional testing to determine the grammar of regulatory sequences that control ion channel gene expression. Using Drosophila melanogaster as a model system, Dr. Atkinson has had substantial success describing how the slowpoke Ca2+-activated K+ channel gene is transcriptionally regulated. It has an extremely complex control region. The Atkinson lab has described control elements that specify the developmental- and tissue-specific expression pattern. Most notable was the identification of control elements that differentially activate one promoter in four different muscle subtypes and the identification of an intronic region that modulates developmental specificity. In the this project Dr. Atkinson will add the Shaker gene to their list of channel genes to be studied. Shaker encodes a voltage-gated K+ channel. Work on slowpoke will also continue. Work on these to genes will enable a comparison of how genes encoding two major classes of K+ channels, the voltage-gated and Ca2+-gated K+ channels, regulate gene expression. In all cases, the focus will be on the regulation of the genes in the nervous system. Transcriptional start sites (~promoters) will be physically mapped and their expression pattern determined using transgenic animals. Because the entire fly is the expression system, the group can study expression in all tissues and organs in their natural developmental context. To identify the control elements the group will make use of the fact that important DNA sequences tend to be conserved over evolutionary time. For each gene, the transcriptional control regions from four different insect species will be sequenced and compared. Small sequences conserved in both sequence and position will be assumed to be control elements. The candidate elements will be tested for function by deleting them from a transgene and then asking how expression had been altered. These methods have been very successful in dissecting the regulation of the slowpoke gene. Furthermore, it will be determined whether the slowpoke and Shaker K+ channels genes show evidence of coordinate regulation; that is, do some of the same combinations of control elements regulate their expression. Because each control element is recognized by a specific transcription factor(s), it will be possible to infer which transcription factors are involved. Drosophila have the same families of ion channel genes found in vertebrates. In addition, most of the important regulatory cascades affecting developmental gene expression were originally discovered in or shown to exist in Drosophila. Therefore, it is expected that the description of the regulation of channel gene expression in Drosophila will also be relevant to the understanding of the same process in other organisms.