MidExDB: A database of DrosophilaCNS midline cell gene expression
© Wheeler et al; licensee BioMed Central Ltd. 2009
Received: 16 June 2009
Accepted: 10 November 2009
Published: 10 November 2009
The Drosophila CNS midline cells are an excellent model system to study neuronal and glial development because of their diversity of cell types and the relative ease in identifying and studying the function of midline-expressed genes. In situ hybridization experiments generated a large dataset of midline gene expression patterns. To help synthesize these data and make them available to the scientific community, we developed a web-accessible database.
MidExDB (Drosophila CNS Midline Gene Expression Database) is comprised of images and data from our in situ hybridization experiments that examined midline gene expression. Multiple search tools are available to allow each type of data to be viewed and compared. Descriptions of each midline cell type and their development are included as background information.
MidExDB integrates large-scale gene expression data with the ability to identify individual cell types providing the foundation for detailed genetic, molecular, and biochemical studies of CNS midline cell neuronal and glial development and function. This information has general relevance for the study of nervous system development in other organisms, and also provides insight into transcriptional regulation.
The neurons and glia that comprise the Drosophila CNS midline cells are an excellent model system to study neurogenesis and gliogenesis [1, 2]. This is due to their highly recognizable location at the midline of the embryo, small number of cells, diversity of cell types, large number of identified genes and associated expression patterns, and the ability to identify individual cell types across embryonic development. In each ganglion, there are ~18 midline neurons including glutamatergic/octopaminergic motorneurons, peptidergic motorneurons, dopaminergic interneurons, and glutamatergic interneurons . There are two molecularly distinct populations of midline glia (MG): the anterior MG (AMG) ensheath the commissural axons that cross the midline and the posterior MG (PMG) have unknown function. Study of the midline cells has been instrumental in studying programmed cell death, the role of the Single-minded (Sim) master regulatory transcription factor protein, neuronal and glial cell fate, neuron-glia interactions, and how diffusible factors control axon guidance. The insect midline cells strongly resemble the floorplate cells that reside at the midline of the vertebrate spinal cord . Both the Drosophila midline cells and vertebrate floorplate cells are important embryonic signaling centers - in Drosophila, the midline cells are a source of signals responsible for axon commissure formation, muscle cell migration, and the formation of the ventral epidermis and mesodermal dorsal median cells.
Construction and content
AP or confocal data is retrieved from MidExDB using one of the search modes on the Home page (described in detail in Utility and discussion section). For example, when using Gene Search to find a gene (Figure 2A), the resulting Images link returns DIC images of whole-mount AP in situ hybridization gene expression data arranged by stage. The Confocal link returns a tabular summary of confocal expression data and a link to the raw data displayed as movies.
Midline-expressed genes newly added to MidExDB.
Summary of midline gene expression data
The 286 midline-expressed genes can be grouped by expression and function. There are 44 genes expressed during the mesectodermal period, 162 at the midline primordium stage, and 198 in mature midline cells (these numbers do not include those genes with potential or uncertain midline expression). Of the genes expressed in mature midline cells, 54 are expressed in MG, 137 in midline neurons, 6 in both, and for 9 genes, the expression cannot be unambiguously assigned. There are 34 genes expressed in midline accessory cells, and 4 present in apoptotic cells dying at the midline. Regarding function, the largest group of genes is concerned with transcription, including 83 members. There are 50 genes listed as cell signaling, and 26 are neural function genes, which encode neurotransmitter biosynthetic enzymes, neuropeptides, membrane transporters, vesicular transporters, and neurotransmitter receptors. The remaining genes are currently partitioned into an additional 21 classes. Previously, we estimated that the 286 midline-expressed genes in this database likely represented >50% of the genes expressed in midline cells (this does not generally include broadly-expressed genes also present in midline cells) .
Utility and discussion
MidExDB has two major search modes: basic data searches and the confocal query tool. The basic data searches return information from AP data and confocal data, while the confocal query tool returns information based on expression in specific cell types. For convenience, the basic data searches (Category, Gene, and Quick) are located on the MidExDB Home page, and the confocal query tool can be accessed on the navigation menu (Figure 2A). Under Category Search, the user can choose to list genes based on: (1) Gene Data (name, symbol or short name, CG#, protein type, or function), (2) Expression Data (AP data), and (3) Confocal Data. Each category is divided into subcategories. For example, the "Gene Data -Midline cell" category can be searched for expression in mesectoderm, midline primordium, mature midline, midline neurons, or MG. The Gene Data category selection results in a list of all genes that is sorted based on selection of a subcategory entry, such as name, protein type, or function. If protein type or function is chosen, then another menu provides a list of proteins and functions to select. For example, selection of "Protein Type" and "bHLH-PAS transcription factor" returns two entries: cycle and sim. Once the list is present, it can be viewed in multiple ways to highlight different aspects of the gene's features and expression. The data is also available in a printable format that can be copied into a database or spreadsheet.
Using Gene Search, the name of a gene is entered and three views are returned: Details, Images, and Confocal. It is also possible to analyze multiple genes together using the Batch Search. The Details view provides an overview of data for that gene. This includes protein type and function, midline cell and midline accessory cell expression data determined from AP data, and an indication if confocal data is available with a link to access that information. There are also links to the FlyBase  and the BDGP gene expression database  entries for the gene. The Images view displays AP histochemically-stained images of midline gene expression from embryonic stages 5 to 17. Below the row of images for each stage is a description of the expression pattern using the controlled vocabulary (Figure 3A-D). The descriptions for stages 9, 10, 11, 13, and 17 also include the results of confocal analysis for those genes analyzed. As an example, an entry for sim is shown for stage 17 in Figure 3A, and DAT at stage 17 in Figure 3B. They are both annotated as expressed in midline neurons based on the AP data, whereas the confocal data indicates that DAT is expressed in one neuron (H-cell), while sim is expressed in a larger set of midline neurons (H-cell sib, ventral unpaired median interneurons 4-6 (iVUMs4-6), median neuroblast progeny (MNB progeny), as well as the median neuroblast (MNB) and AMG. The Confocal view (Figure 2B) provides a text-based summary of confocal expression data for the selected gene. The confocal data is divided into stages 9, 10, 11, 13, 17 with each corresponding cell type listed.
At the bottom of the MidExDB home page are 6 Quick Searches for AP data or confocal data (Figure 2A). The quick searches can be used to retrieve a list of all genes with either AP data or confocal data. There is also a quick search function for retrieving all genes expressed in MG or midline neurons. From the retrieved lists, clicking on the gene symbol will open the Details page for that gene.
The second major search mode, the Confocal Query Tool allows the user to customize a query to identify genes expressed in specific cell types at defined developmental stages (Figure 2C). Each cell type is listed for each of the 5 developmental stages in which confocal data is present. Complex queries using the Boolean operators AND, OR, and NOT are possible. As an example (Figure 2C), the database can be searched for genes expressed at: stage 13 in the MP1 neurons AND H-cell sib but NOT H-cell. Only 1 gene, fork head (fkh) appears. Its expression at each stage of development is listed in tabular form (Figure 2C). Lists of genes obtained using the Confocal Query Tool can be printed or exported to the Category Search to view additional data (for example, protein type) for each gene in the list.
Descriptive background information and protocols
The Protocols section in the Information menu provides PDFs of detailed experimental protocols that were used to generate the data present in MidExDB. Antibody staining and in situ hybridization protocols are described for both fluorescence and AP detection, including the use of Tyramide Signal Amplification (TSA).
The goal of MidExDB is to disseminate information about the Drosophila CNS midline cells. MidExDB is able to search and integrate a large-scale midline gene expression dataset. The database provides a useful foundation for studying CNS midline development and function, particularly the underlying regulatory circuitry.
Availability and requirements
The authors would like to thank Joseph Kearney, Amaris Guardiola, and Joseph Pearson for data entry and useful advice. We are also grateful to Hínár Polczer for his help with software and server maintenance, and Bob Duronio for helpful comments on the manuscript. This work was supported by NIH grant R37 RD25251 to STC, an NRSA postdoctoral fellowship to SRW, and UNC Developmental Biology NIH training grant support to SBS.
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