Casein-kinase-II-dependent phosphorylation of PPARgamma provokes CRM1-mediated shuttling of PPARgamma from the nucleus to the cytosol - PubMed (original) (raw)

. 2010 Jan 15;123(Pt 2):192-201.

doi: 10.1242/jcs.055475. Epub 2009 Dec 21.

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Casein-kinase-II-dependent phosphorylation of PPARgamma provokes CRM1-mediated shuttling of PPARgamma from the nucleus to the cytosol

Andreas von Knethen et al. J Cell Sci. 2010.

Abstract

PPARgamma exerts significant anti-inflammatory signaling properties in monocytes and macrophages, which are affected by its intracellular localization. Based on our previous report, which showed that cytosolic localization of PPARgamma attenuates PKCalpha signaling in macrophages, we elucidated the molecular mechanisms provoking cytosolic PPARgamma localization. Using the DsRed-tagged PPARgamma deletion constructs PPARgamma1 Delta1-31 and PPARgamma1 Delta407-475, we observed an exclusive nuclear PPARgamma1 Delta1-31 localization in transfected HEK293 cells, whereas PPARgamma1 Delta407-475 did not alter its cytosolic or nuclear localization. The casein kinase II (CK-II) inhibitor 5,6-dichloro-1-beta-D-ribofuranosyl benzimidazole (DRB) prevented cytosolic PPARgamma localization. Mutation of two possible CK-II phosphorylation sites at serine 16 and serine 21 of PPARgamma into alanine (PPARgamma S16A/S21A) inhibited cytosolic PPARgamma localization. Moreover, a PPARgamma S16E/S21E mutant that mimicks constitutive phosphorylation of residues 16 and 21, predominantly resides in the cytosol. The CRM1 inhibitor leptomycin B abolished cytosolic PPARgamma localization, suggesting that this is a CRM1-dependent export process. CRM1-mediated PPARgamma export requires Ran and phosphorylated RanBP3. Finally, co-immunoprecipitation studies demonstrated that DRB blocks PPARgamma binding to CRM1, whereas PD98059 inhibits RanBP3 binding to CRM1 and concomitant shuttling from nucleus to cytosol, but does not alter PPARgamma binding to CRM1. We conclude that CK-II-dependent PPARgamma phosphorylation at Ser16 and Ser21 is necessary for CRM1/Ran/RanBP3-mediated nucleocytoplasmic translocation of PPARgamma.

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