Evidence for importin alpha independent nuclear translocation of glucocorticoid receptors in Xenopus laevis oocytes

Abstract

The glucocorticoid receptor (GR) is a ligand-dependent transcription factor which resides in the cytoplasm as a complex with a number of molecular chaperones. Upon hormonal stimulation, the GR is translocated into the nucleus where it modulates transcription by binding to specific target DNA sequences. GR contains a classical basic nuclear localization signal (NLS) and a second, only poorly characterized NLS. Though the participation of the importin alpha/importin beta pathway in nuclear import of GR has been postulated, relatively little is known about its dynamics and mechanisms. We analyzed the hormone-driven nuclear import of rat GR expressed in Xenopus laevis oocytes by atomic force microscopy (AFM) and western blot analysis. AFM imaging revealed accumulation of macromolecules matching the size of GR at the nuclear envelope beginning 5 min after glucocorticoid hormone injection. In parallel, western blot analysis showed accumulation of GR over the same time scale after glucocorticoid hormone stimulation. Nuclear GR import did not trigger redistribution of importin alpha or importin beta. We conclude that hormone-driven nuclear import of rat GR expressed in Xenopus oocytes is a process with limited capacity and involves mechanisms different from the importin alpha/importin beta pathway. This represents a restriction of nuclear import downstream of import signals.