© The Rockefeller University Press,
0021-9525/1997//523 $5.00
The Journal of Cell Biology, Volume 137, Number 3,
, 1997 523-538
Subnuclear Trafficking of Glucocorticoid Receptors In Vitro: Chromatin Recycling and Nuclear Export
Jun Yang,
Jimin Liu, and
Donald B. DeFranco
Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania 15260
We have used digitonin-permeabilized cells to examine in vitro nuclear export of glucocorticoid receptors (GRs). In situ biochemical extractions in this system revealed a distinct subnuclear compartment, which collects GRs that have been released from chromatin and serves as a nuclear export staging area. Unliganded nuclear GRs within this compartment are not restricted in their subnuclear trafficking as they have the capacity to recycle to chromatin upon rebinding hormone. Thus, GRs that release from chromatin do not require transit through the cytoplasm to regain functionality. In addition, chromatin-released receptors export from nuclei of permeabilized cells in an ATP- and cytosol-independent process that is stimulated by sodium molybdate, other group VI-A transition metal oxyanions, and some tyrosine phosphatase inhibitors. The stimulation of in vitro nuclear export by these compounds is not unique to GR, but is restricted to other proteins such as the 70- and 90-kD heat shock proteins, hsp70 and hsp90, respectively, and heterogeneous nuclear RNP (hnRNP) A1. Under analogous conditions, the 56-kD heat shock protein, hsp56, and hnRNP C do not export from nuclei of permeabilized cells. If tyrosine kinase inhibitors genistein and tyrphostin AG126 are included to prevent increased tyrosine phosphorylation, in vitro nuclear export of GR is inhibited. Thus, our results are consistent with the involvement of a phosphotyrosine system in the general regulation of nuclear protein export, even for proteins such as GR and hnRNP A1 that use distinct nuclear export pathways.
1. Abbreviations used in this paper: DAPI, 4',6-diamidino-2-phenylindole; GR, glucocorticoid receptor; hnRNP, heterogeneous nuclear RNP; IIF, indirect immunofluorescence; NES, nuclear export signal sequence; NLS, nuclear localization signal sequence; NPC, nuclear pore complex.
Please address all correspondence to Donald B. DeFranco, Department of Biological Sciences, University of Pittsburgh, Pittsburgh, PA 15260. Tel.: (412) 624-4259. Fax: (412) 624-4759. e-mail: dod1{at}vms.cis.pitt.edu

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