Interaction with MEK Causes Nuclear Export and Downregulation of Peroxisome Proliferator-Activated Receptor γ

The peroxisome proliferator-activated receptor γ (PPARγ) is a nuclear receptor that participates in the regulation of a large number of cellular processes including differentiation, immune response, and metabolism (31). The activity of PPARγ is normally induced by binding of specific ligands that activate its genomic transcriptional activity and thus initiate the expression of several effector genes. In addition, as a central signaling component, the activity of PPARγ is well regulated and can be inhibited under various cellular conditions such as stimulation of cells with growth factors and protein kinase C activators (24). Among the molecular mechanisms that prevent the activity of PPARγ upon these conditions is the phosphorylation of Ser84/Ser112 within PPARγ1/PPARγ2 by mitogen-activated protein kinases (MAPKs)/extracellular-signal regulated kinases 1 and 2 (ERK1/2) (14, 22). Stress stimuli (11) and proinflammatory mediators (30) also prevent PPARγ activation, but this is mediated mainly by other MAPK cascades including c-Jun N-terminal kinase and p38MAPK, which seem to induce phosphorylation of Ser84/112 as well. The inhibition of PPARγ by growth factors and gamma interferon are in accordance with the antiproliferative and anti-inflammatory role of PPARγ. Interestingly the MAPK cascade of ERK5 was reported to interact with PPARγ, but unlike the other MAPKs, this direct interaction induces activation rather than inhibition of PPARγ transcriptional activity (4). Finally, it was also shown that PPARγ ligands can trigger an activation of the ERK cascade (19, 37); however, the exact role of this activation is not clear as yet. One important parameter that participates in the regulation and function of PPARγ as well as MAPK signaling is their subcellular localization (27, 42). Regarding the ERK cascade, it was shown that both ERKs and MEKs are localized in the cytosol of resting cells and that they translocate into the nucleus upon cellular stimulation (46). However, while ERKs remain in the nucleus of the stimulated cells for up to 180 min, MEKs are rapidly exported out of the nucleus due to their nuclear export signals (NES) (23). In addition, slow cytonuclear shuttling of inactive MEKs can occur without extracellular stimulation (18) and may assist export of inactive ERKs from the nucleus (1). Unlike the ERK components, little is known about the regulation of the intracellular distribution of PPARs. PPARγ was shown in several studies to localize mainly in the nucleus (6, 20). However, evidence for a significant cytosolic localization upon ligand binding has been presented as well (35, 39, 42). Further indications for extranuclear localization may be deduced from the reported relationships with cytosolic/membranal proteins such as HSP90 (36) and caveolin-1 (10). Therefore, although PPARγ functions primarily in the nucleus, it can redistribute to different compartments under distinct physiological conditions. However, no NES or a definite “shuttle” protein for PPARγ has been described, and therefore, the mechanism that allows the redistributions is not clear. Interestingly, the cytoplasmic retention sequence/common docking domain (CRS/CD) (32, 38) of ERK exhibits significant sequence similarity to the AF2 motif of PPARγ (9). Since CRS/CD participates in ERK-MEK interaction by docking to MEKs' D domain (38), it is possible that the CRS-like motif in PPARγ may mediate a selective interaction with MEKs. Here we report that a physical association between MEK1 and PPARγ does exist, and we show that this interaction is important for the subcellular localization and transcriptional activity of PPARγ. Our data support a model in which PPARγ is localized in the nucleus of resting cells. Mitogenic stimulation of the cells causes interaction of active nuclear MEKs with PPARγ, and this is followed by a rapid nuclear export of the complex, which is mediated by the NES of MEKs. This nuclear export reduces the transcriptional activity of PPARγ and allows it to interact with cytosolic and membranal components. Therefore, we show here a novel role for MEKs, beside the regulation of ERKs, which is determination of the subcellular localization of other nuclear proteins to modulate their activities.