MicroRNA-mediated regulatory control of class specific dendrite morphogenesis in Drosophila
Date
2014-10-07
Authors
Sivakumar, Shruthi
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Abstract
Dendrites function as the primary sites of synaptic and sensory input and integration in the developing nervous system. Moreover, numerous cognitive disorders such as Autism, Rett’s syndrome, Angelman’s syndrome and Schizophrenia are associated with dendritic abnormalities including development and maintenance (Kaufmann and Moser, 2000). MicroRNAs (miRNAs) have recently emerged as critical post-transcriptional regulators of gene expression in numerous tissues, particularly the nervous system, with functions in specification of neuronal development, formation and maintenance of dendritic fields, neuronal asymmetry, axonal and dendrite targeting and local translation at synapses based on neuronal activity. Despite the aforementioned advances, the precise role of miRNA-mediated regulation of neuronal morphogenesis and development of class specific dendrite arborization patterns remains largely unknown. The peripheral nervous system (PNS) of Drosophila melanogaster provides an excellent system in which to elucidate the molecular mechanisms governing class specific dendrite morphogenesis and homeostasis using dendritic arborization (da) sensory neurons as a model system. Based on the central hypothesis that miRNA-mediated post-transcriptional regulation is required for establishing and/or maintaining distinct class specific dendritic morphologies, the studies described herein focus on large-scale, comparative morphological analyses of the roles of miRNAs in mediating class-specific da sensory neuron dendrite development via miRNome-directed gain-of-function genetic screens and corresponding loss-of-function mutant analyses. To assess the biological significance of these deregulated miRNA phenotypes, it is important to examine whether these gain-of-function effects are relevant to native endogenous gene function. To directly address this question, we elected to analyze a particular miRNA cluster, composed of miR-12, miR-283 and miR-304 (miR-12/283/304 cluster) via both mutant and overexpression phenotypic analyses. Expression analyses reveal that miR-12 and miR-304 are differentially enriched in complex class III and IV da neurons relative to the morphologically simple class I neurons. MARCM mutant analyses implicate the miR-12/283/304 cluster in cell autonomously regulating higher order dendritic branching, particularly the formation of actin-rich dendritic filopodia, as well as in promoting overall dendritic growth. To dissect the individual contributions of miRNA cluster components, we conducted overexpression studies which revealed differential roles for these miRNAs in mediating class-specific dendritic diversity. Overexpression studies indicate that miR-12, and to a lesser extent miR-304, act in a similar direction to promote dendritic growth and branching, especially actin-rich filopodia, whereas miR-283 acts to restrict these processes such that with full cluster overexpression there is an intermediate phenotype reflective of the opposing functional roles of these miRNAs in mediating class III dendrite morphogenesis. Intriguingly, while the same regulatory effects of these miRNAs were observed in class IV neurons, we found that overexpression of miR-12 in these neurons resulted in the de novo formation of actin rich filopodia and produced a shift in class IV neurons towards class III arborization characteristics indicating that regulated expression of members of this miRNA cluster is important for promoting class-specific dendritic diversity. Collectively, these studies provide both global insight and a robust foundation on the importance of miRNAs in mediating class specific dendrite morphogenesis and will potentially shed novel light on the molecular mechanisms via which miRNA regulation underscores such biologically relevant events as learning and memory, as well as nervous system disease pathologies.
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Keywords
MicroRNA, Morphology, Dendrite, Neurons, MiR-12/283/304, Cytoskeleton