Principal InvestigatorTitle: “Developing drivers for neuron type-specific gene expression”
BRAIN Category: (RFA MH-14-216)
Dr. Hobert and colleagues will create a highly selective technology for experimentally manipulating genes in neurons, by tapping into the regulatory machinery of individual cell types.
Driver lines that direct Cre protein to specific neuron types have proven to be invaluable tools to not only visualize specific neuron types but also to manipulate their activity through the Cre- mediated activation of optogenetic probes or to assess gene function by Cre-mediated gene knockout. Most Cre driver lines, such as BAC-based Cre drivers or knock-ins of Cre into specific loci, monitor the complete expression pattern of entire genetic loci. However, very few genes are exclusively expressed in very small populations of specific neuron types and this lack of cellular specificity limits the use of these driver lines. W propose here to develop transgenic mouse driver lines that direct Cre expression to very restricted numbers of neuronal cell types in different regions of the mouse brain, thereby providing tools to precisely map their function and molecular composition. To achieve this aim, we aim to test the hypothesis – built from our past work in the nematode C.elegans – that the cis-regulatory control elements of the mouse loci that encode the vesicular transporters for the four main neurotransmitter systems in the vertebrate central nervous system, glutamate and -aminobutyric acid (GABA) and acetylcholine (ACh) are composed of a modular assembly of individual, highly cell type-specific cis-regulatory elements. We will experimentally test the hypothesis that the expression of individual, isolated cis-regulatory elements may subdivide cholinergic, glutamatergic and GABAergic domains into restricted and perhaps novel domains of the mouse central nervous system and thereby constitute reproducible and highly specific drivers for directing the expression of genes that allow the genetic manipulation of neurons and neuronal circuits. This cis-regulatory dissection approach may solve the specificity problem of most currently available driver lines that are unable to exclusively target restricted numbers of cells.
Public Health Relevance Statement
One of the central tenets of molecular and cellulear neuroscience is that if one wants understand how the brain develops and functions, it is of critical importance to be able to visualize and manipulate the activity of the individual building blocks of the brain. In theory, individual cells in the nervous system can be visualzed, activated or inhibited using the expression of specific, well characterized genes. However, our ability to direct the expression of such genes to specific neuron types, specific tools are required that are currently available only to a very limited extent. We propose here to define pieces of DNA that are able to direct expression of any gene of interest to very specific types of neurons in the mouse brain which would then allow to visualize and manipulate their activity.
NIH Spending Category
Biotechnology; Genetics; Mental Health; Neurosciences
Acetylcholine; acetylcholine transporter; Adult; Aminobutyric Acids; base; Binding Sites; Brain; Caenorhabditis elegans; Cell Count; cell type; Cells; cholinergic; Dissection; DNA; Elements; Enzymes; Future; Gene Expression; gene function; Generations; Genes; Genetic; genetic manipulation; Genomics; Glutamates; Individual; insight; Integrase; interest; knockout gene; LacZ Genes; Maps; Mediating; Molecular; Monitor; mouse genome; Mus; Mutate; Nature; Nematoda; Nervous system structure; Neuraxis; Neurons; Neurosciences; Neurotransmitters; novel; Nucleic Acid Regulatory Sequences; optogenetics; Pattern; Phenotype; Population; Positioning Attribute; Proteins; public health relevance; Regulatory Element; Reporter; Specificity; System; Testing; theories; tool; transcription factor; Transgenic Mice; Transgenic Organisms; Work