NUCLEOCYTOPLASMIC TRANSPORT--NEW FACTORS AND PATHWAYS

Mount Sinai School Of Medicine Of Nyu, New York NY

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Abstract

Various classes of proteins and nucleic acids are imported into and exported out of the cell nucleus through separate pathways, defined by independent sets of carrier molecules and docking sites ad the nuclear pore complexes. For some transported species the components of their transport machineries are partially or totally unknown. Our aim is to identify and characterize new nucleocytoplasmic transport factors, based on two complementary strategies. The first approach involves obtaining the full DNA sequences of potential transport factors whose existence is inferred from the presence in the GenBank EST database of shot, partial cDNA sequences, showing significant homology with known transport factors. The second strategy is based on the fact that all known transport factors dock temporarily, during passage across the nuclear pore, to proteins at the nuclear pore complexes. One of these docking site proteins, Nup98, which we isolated and shown to be implicated in multiple transport pathways, will be used for retention and isolation of new transport factors from the nucleoplasm. The proteins isolated by these two approaches will be characterized in terms of full cDNA sequence, subcellular localization, and level of expression in various tissues and developmental stages. Most importantly, the proteins interacting with these transport factors will be identified by co-immunoprecipitation, screening of expression libraries, and two-hybrid assays. The ensemble of these data, and especially the set of interacting molecules, will shed light on the specific functions of these proteins as transport factors. The relevance of these studies for human health is linked to the fact that a vast number of proteins and nucleic acids cannot exert their function unless they are imported in or exported out of the nucleus in a precise manner, and in fact there are known cases when tumors arise due to defects in nucleocytoplasmic transport pathways are used by viruses, knowing in detail these mechanisms could also offer opportunities for new anti-viral therapies and for improved gene therapy vectors.