The World Health Organization estimates that nearly 500 million people suffer from various forms of mental illness worldwide, and trending data predicts that by the year 2030 depression will be the single highest contributor to burden of disease in the world. Many mental health disorders, including depression, are associated with a change or disruption of dendrites and synaptic contacts, which are established by dynamic processes within individual neurons controlling morphology during development. Some of the best-studied regulators of neuron morphology are the Rho GTPases, namely Cdc42 (Cell division cycle 42) and Rac1 (Ras- related C3 botulinum toxin substrate). These proteins have a remarkable similarity in their amino acid sequence and apparent functional roles. Previous work extensively used over-expression of wild type and mutant forms of these proteins and lead to a popular notion that the two have very similar influences on cytoskeletal dynamics. However, there is an apparent functional divergence left to be explored. Both are physiologically essential, yet how they differ in establishing neuron morphology is unclear. Downstream to both Cdc42 and Rac1 is a group of effectors that appear to be instrumental in neurite development that are known as CRIB proteins. Determination of when and where Cdc42 and Rac1 are activated, and how the timing and location of their activation contribute to their downstream effects within central neurons will be important for understanding molecular control of neuronal morphology and synaptic connectivity. These ancient signaling mechanisms are fundamental to all eukaryotes, and genetic tools available in Drosophila melanogaster will facilitate their study while also giving insight into mammalian nervous systems. My central hypothesis is that Cdc42 and Rac1 are not functionally redundant proteins, that they indeed have unique signaling properties, and that timing and location of interactions each has with downstream CRIB effectors is crucial for proper morphological development in the Drosophila anterior corner cell (aCC) neuron. The objectives of this application are to: (1) establish the activation pattern of Rac1 during Drosophila aCC neuron development using a FRET aProbe construct, complementing that of Cdc42 demonstrated previously in the sponsor's laboratory;(2) see where and when Cdc42 and Rac1 bind with CRIB protein partners in vivo using both ratiometric and lifetime FRET imaging approaches, and (3) determine through loss of function studies how CRIB protein partners contribute to neuronal morphology. By determining for Rho GTPases both their signaling pathways and their functional contributions to the formation of neuronal compartments in living neurons, the proposed studies will provide important information on regulation of neuronal connections. A compelling aspect of this work will be visualizing how the neuron integrates converging signaling pathways to grow neurites and form synapses. By understanding the molecular basis of these processes, medicine will gain novel therapeutic targets to combat the pathophysiologies that afflict those with debilitating neuropsychological conditions.
The World Health Organization estimates that nearly 500 million people suffer from various forms of mental illness worldwide, and trending data predicts that, by 2030, depression will be the single highest contributor to burden of disease in the world. As many mental disorders are associated with the loss of synaptic contacts between neurons, this project will provide a better understanding of how normal synapses are formed by protein interactions in the neuron during development. In doing so, it will aid in the development of targeted treatments for mental illnesses so that we may reduce the social and economic burden of this growing epidemic.
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|Sharifai, Nima; Samarajeewa, Hasitha; Kamiyama, Daichi et al. (2014) Imaging dynamic molecular signaling by the Cdc42 GTPase within the developing CNS. PLoS One 9:e88870|