Nup50, a nucleoplasmically oriented nucleoporin with a role in nuclear protein export

We present here a detailed analysis of a rat polypeptide termed Nup50 (form erly NPAP60) that was previously found to be associated with the nuclear po re complex (F. Fan et al., Genomics 40:444-453, 1997). We have found that N up50 (and/or a related 70-kDa polypeptide) is present in numerous rat cells and tissues. By immunofluorescence microscopy, Nup50 was found to be highl y concentrated at the nuclear envelope of rat liver nuclei, whereas in cult ured NRK cells it also is abundant in intranuclear regions. On the basis of immunogold electron microscopy of both rat liver nuclear envelopes and NRK cells, we determined that Nup50 is specifically localized in the nucleopla smic fibrils of the pore complex. Microinjection of anti-Nup50 antibodies i nto the nucleus of NRK cells resulted in strong inhibition of nuclear expor t of a protein containing a leucine-rich nuclear export sequence, whereas n uclear import of a protein containing a classical nuclear localization sequ ence was unaffected. Correspondingly, CRM1, the export receptor for leucine -rich export sequences, directly bound to a fragment of Nup50 in vitro, whe reas several other import and export receptors did not significantly intera ct with this fragment. Taken together, our data indicate that Nup50 has a d irect role in nuclear protein export and probably serves as a binding site on the nuclear side of the pore complex for export receptor-cargo complexes .