Structure-function analysis of RanBP2 in the neuroretina

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DESCRIPTION (provided by applicant): Protein trafficking is a fundamental biological process across all eukaryotic cells. A central question in cell physiology is how the cell manages to accommodate the trafficking of a plethora of cargoes with a very limited number of motor proteins. A wide variety of modulators mediate the intracellular transport of a plethora of endogenous cargoes between subcellular compartments, and pathogens often hijack components of this transport machinery. Emerging evidence also supports this process is vital for triggering defined cellular events such as the onset of mitosis and cell death. Protein kinesis assumes special relevance in neuronal systems in light of the exquisite compartmentalization and polarization of subcellular compartments and organelles across neurons, and significant burden imposed on neurons for the long distances cargoes often need to travel to reach their final destinations. A large and growing number of genetic lesions in trafficking components and interacting cargoes, lead to a host of neuropathies and allied maladies in the human. In this proposal, the neuroretina is the experimental system of choice to study protein kinesis because it is a diverse but extremely well defined biological system highly amenable to manipulations in vitro and in vivo. In addition, there are a large number of neurodegenerative retinal dystrophies, which affect directly and indirectly numerous facets of processes mediating protein trafficking. The focus of this proposal is to understand the role of an unique and large scaffold vertebrate protein, Ran-binding protein 2 (RanBP2/Nup358), in mediating and integrating key steps of intracellular trafficking processes and protein biogenesis, and implications of these processes in retinal cell physiology and etiology of pathological states. Past work has implicated RanBP2 in mediating in vivo the functional production of light-receptors and nuclear protein import. We have mapped and characterized domains of RanBP2, identified associated molecular partners and determined the spatial localization of these among and within retinal neurons. Moreover, a large number of molecular, biochemical, immunological and genetic tools were developed. These, in combination with other interdisciplinary approaches, will now be used to probe in vitro and in vivo specific and emerging trafficking pathways mediated by RanBP2 within retinal neurons, to identify novel partners and cargoes, regulatory mechanisms modulating the assembly of RanBP2 complexes, and to dissect further the molecular basis of neuronal-restricted transport pathways mediated by RanBP2. Finally, we will investigate the physiological effects of these components and processes in the pathogenesis of retinal dystrophies.