Transcriptional Regulation Of Neuronal Differentiation In The Drosophila Central Nervous System

The central nervous system is the most complex and highly organized tissue in animals; composed of thousands of neurites connected in specific and highly reproducible ways. My thesis research has focused on the generation of neuronal diversity: specifically how neurons adopt individual, often unique, identities. Work in many labs has revealed that a large set of transcription factors act in combinatorial manner to specify the fate of individual neurons or small groups of neurons. However, in most cases, it remains unclear how individual or specific combinations of transcription factors directly control the terminal differentiation of neurons via the regulation of different genes, such as neurotransmitters. My thesis work has focused on the identification and characterization of new members of the combinatorial code of transcription factor and on initial attempts to link these transcription factors to the expression and activity of genes that contribute directly to neuronal differentiation. In chapter 2, I describe the identification and characterization of Dbx, a homedomain-containing transcription factor, expressed in a mixture of progenitor cells and a subset of GABAergic interneurons. I show that Dbx is expressed in many interneurons that are sibling to motor neurons, and that Dbx is required to promote the development of these interneurons via cross-repressive interactions with Eve and Hb9, which are expressed in the sibling motor neurons. In chapter 3, I detail the identification of FoxD, a transcription factor that is positively regulated by the homeodomain-containing transcription factor Hb9 in the Drosophila CNS. FoxD is expressed in a subset of Hb9 positive neurons and also in all octopaminergic neurons in the Drosophila embryonic CNS. I have identified the enhancers that drive expression in these neurons and have recently generated two mutant alleles of foxD. Loss of foxD appears to result in hyperactivity, which is most pronounced in males. As octopamine is the fly equivalent of norepinephrine, these results suggest that FoxD may function in specific cells to regulate the synthesis and release of octopmaine. Thus, my thesis has identified two members of the combinatorial code of transcription factors that govern neuronal identity. In addition, it has begun to place the functions of these genes within the genetic regulatory hierarchy of this code and started to link the function of individual transcription factors to the regulation of terminal differentiation genes and animal behavior.

This first book to cover neural development, neuronal survival and function on the genetic level outlines promising approaches for novel therapeutic strategies in fighting neurodegenerative disorders, such as Alzheimer's disease. Focusing on transcription factors, the text is clearly divided into three sections devoted to transcriptional control of neural development, brain function and transcriptional dysregulation induced neurological diseases. With a chapter written by Nobel laureate Eric Kandel, this is essential reading for neurobiologists, geneticists, biochemists, cell biologists, neurochemists and molecular biologists.

Abstract: During Drosophila embryogenesis, tight coordination between cell proliferation and terminal differentiation is required to ensure the proper formation of the nervous system. However, little is known regarding the mechanism coordinating cell cycle proliferation and terminal differentiation. The main goal of my research was to analyze the transcriptional regulation of the cyclin-dependent kinase inhibitor (CKI) dacapo (dap) gene expression during Drosophila melanogaster neurogenesis. dap is the only identified G1 CKI in Drosophila and is a homolog of p27kip. I found dap expression to be regulated by a complex array of tissue-specific cis-regulatory elements. prospero (pros), a pan-neural transcription factor, regulates dap expression in the embryonic nervous system. Furthermore, Pros and DmcycE, the Drosophila homolog of cyclin E, function cooperatively in regulating the expression of both dap and the neuronal differentiation marker-Even-skipped (Eve). A second goal of my research was to analyze the role of Pros in the regulation of mitotic activity and differentiation. Evidence is presented that cell cycle regulatory genes are downstream targets of Pros in regulating mitotic activity. In addition, Pros interacts with cell cycle regulatory genes to regulate the expression of neuronal differentiation markers in a lineage specific pattern.

Determinants of Neuronal Identity brings together studies of a wide range of vertebrate and invertebrate organisms that highlight the determinants of neuronal identity. Emphasis of this book is on how neurons are generated; how their developmental identities are specified; and to what degree those identities can be subsequently modified to meet the changing needs of the organism. This book also considers various techniques used in the analysis of different organisms. This volume is comprised of 15 chapters; the first of which introduces the reader to the specification of neuronal identity in C ...

The last decades have witnessed a radical change in our views on central nervous system damage and repair. This change is not only due to the emergence of new powerful tools for the analysis of the brain and its reactions to insults, but it also reflects a conceptual change in the way we approach these problems. As an illustration to this development, it is instructive to go back to the proceedings of a meeting at the NIH in 1955 edited by William F. Windle, which summarizes the disillusioned and pessimistic view on CNS regeneration prevailing at the time. While this generation of researchers were well aware of the issues at stake, they felt they had reached the end of the road; the approaches they had pursued had got stuck and the tools available could not take them any further. I can very well imagine that the participants, most of them leaders in the field, left that conference feeling they had heard their field being sentenced to death.

Neurons are a diverse group of cells, with their unique morphology often linked to specific functional requirements. The intricate morphology of neurons necessitates a multilayered developmental process, with regulation occurring at the transcriptional, post-transcriptional and post-translational levels. RNA-binding proteins (RBPs) that mediate post-transcriptional regulation can function at all stages of the lifecycle of a transcript, allowing for rapid and localized control over gene expression. These regulatory features are particularly important in cells as expansive and morphologically complex as neurons. The Drosophila larval class IV dendritic arborization (da) neurons provide an ideal system for studying the role of RBPs in neuron development. These highly branched sensory neurons completely and non-redundantly tile the larval body wall. Here, we analyze the role of the RBP Found in neurons (Fne) in regulating the space-filling dendrite growth that is characteristic of these neurons. Our results indicate that Fne regulates multiple transcripts and in doing so, is able to modulate both processes intrinsic to the neuron as well as interactions between the neuron and the surrounding environment. By regulating multiple, functionally-related transcripts in tandem, Fne is able to act as a central coordinator of neuronal morphology. This synchronized regulation is likely to be an important feature of many other RBPs that have been implicated in neuronal morphogenesis. Furthermore, our results reveal that the molecular mechanisms underlying the role of Fne in class IV da neurons are distinct from its recently revealed functions in the central nervous system. The differences in Fne function in various types of neurons, and therefore its participation in distinct regulatory processes, could be a feature that is common to many RBPs and may contribute to the vast morphological diversity observed in neurons.

The fruitfly Drosophila melanogaster is an ideal model system to study processes of the central nervous system This book provides an overview of some major facets of recent research on Drosophila brain development.

Abstract: The goal of developmental neurobiology is to understand how a complex nervous system is wired. During development of the central nervous system (CNS) neural connections are assembled in a highly stereotyped fashion. How do axons find their targets with such accuracy? We know that axon migration is direct by attractive and repulsive guidance cues located in the extracellular environment. While many guidance molecules have been identified, we are only just beginning to understand the mechanisms of axon guidance. In order to identify additional genes involved in axon guidance and CNS development we performed a misexpression screen. Using P-elements and the UAS/GAL4 system, transcription of endogenous genes was induced in the embryonic CNS. Misexpression phenotypes were then identified immunohistochemically with two monoclonal antibodies: BP102, a general axon marker, and 1D4, which labels a subset of axon pathways. Over 4100 individual P-element insertion lines were screened. Twenty-five insertions corresponding to 18 genes resulted in misexpression phenotypes. Genes involved in axon guidance, embryonic patterning, and cell cycle regulation were identified. Several transcription factors that have not been previously implicated in CNS development were isolated and characterized as well. The identification of these transcription factors is intriguing since little is known about the transcriptional regulation of axon guidance genes. Additionally, we have studied the regulation of the previously identified guidance molecule Commissureless (Comm). Comm is necessary for proper axon guidance at the CNS midline of the Drosophila embryo. In the absence of Comm, commissural axons fail to cross the midline and instead make ispilateral projections on their respective sides of the midline. Using mosaic analysis, we have identified a cell autonomous neuronal requirement for Comm. Clones containing mutant alleles of comm formed commissural projections at a statistically significant reduced frequency when compared to wild type clones. This result suggests that regulation of Comm expression in neurons is critical for Comm's function in axon guidance at the CNS midline. These studies have both advanced the understanding of the regulation of Comm, and have identified new potential regulators of guidance molecules.

Drosophila--the scientist's favorite laboratory organism--is a key subject for studying genetics, development, and behavior. The second edition of this popular hands-on guide brings the methodology up-to-date, with contributions from the leading researchers in the field. Chapters include the elements of Drosophila biology and genetics; mutagenesis; transposons--gene tagging and mutagenesis; chromosome mechanics: the genetic manipulation of aneuploid stocks; enhancer traps; looking at embryos; immunolabelling of Drosophila; population and ecological genetics; behavior, learning, and memory; cell culture; and preparation of nucleic acids.