Nuclear transport serves a critical regulatory function by modulating the activities of transcription factors, nuclear kinases, steroid hormone receptors and replication factors. The nuclear pore complex (NPC) mediates the transport of mRNA and proteins across the nuclear envelope. Previously, we described a family of phosphorylated glycoproteins of the NPC whose posttranslational modifications appear to be associated with assembly and disassembly of nuclear pores. We have molecularly cloned and characterized the human O-linked GlcNAc transferase responsible for glycosylating nuclear pore proteins. Using both the mouse and Caenorhabditis elegans homologs of the enzyme we are attempting to genetically manipulate the enzyme in an intact organism. Using digitonin permeabilized cultured cells, we have studied nuclear and nucleolar transport of fluorescent conjugates bearing the appropriate localization signal. In addition to the well characterized GTP-dependent nuclear transport observed in permeabilized cells, we detected a mode of nuclear transport that was GTP-independent at elevated cytoplasmic calcium concentrations. Nuclear transport under these conditions was blocked by calmodulin inhibitors. Recombinant calmodulin fully restored ATP-dependent nuclear transport in the absence of cytosol. Calmodulin-dependent transport was inhibited by wheat germ agglutinin suggesting that transport proceeded through nuclear pores. We propose that release of intracellular calcium stores upon cell activation inhibits GTP- dependent nuclear transport; the elevated cytosolic calcium then acts through calmodulin to stimulate the novel GTP-independent mode of transport. This provides a direct link between signal transduction and the modulation of nuclear transport.
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