JAB1 (Jun activation domain-binding protein-1) has previously been described as a coactivator of AP1 transcription factor. We show here, by yeast and mammalian two-hybrid analyses and by pull-down experiments, that JAB1 also interacts with both the progesterone receptor (PR) and the steroid receptor coactivator 1 (SRC-1) and that it stabilizes PR-SRC-1 complexes. We also show that JAB1 potentiates the activity of a variety of transcription factors known to associate with SRC-1 (nuclear receptors, activator protein-1, and nuclear factor B). This occurs without any modification of PR or SRC-1 concentration. JAB1 is a subunit of a large multiprotein complex that has been called the COP9 signalosome. The latter is present in plant and animal cells and has been shown to be involved in a variety of cellular mechanisms including transcription regulation, cell cycle control, and phosphorylation cascades. We now show that it is also involved in the mechanisms of action of nuclear receptors and of their coactivators.JAB1 (Jun activation domain-binding protein-1) was initially isolated by a two-hybrid screen (1) using the c-Jun Nterminal activation domain as a bait. It was shown that JAB1 potentiates target gene transcription activation by AP1 proteins (especially c-Jun and JunD) (1). This initial study also pointed to the sequence homology between JAB1 and the yeast protein Pad-1. The latter has been demonstrated to be necessary for the transcriptional activity of Pap-1, the yeast equivalent of c-Jun (1, 2). Furthermore it was shown that JAB1 could functionally replace Pad-1 in yeast. Thus JAB1 and Pad-1 appeared to play similar roles in mammalian and yeast cells, respectively. However, recent studies have uncovered the true human homolog of Pad-1, which has been called POH-1. Both Pad-1 and POH-1 are components of the 26 S proteasome (3, 4).On the contrary JAB1 is not found in the proteasome but has very recently been shown to be a subunit of a different multiprotein complex that has been called the signalosome (or COP9 signalosome). This complex contains several proteins with sequence homologies to proteins present in the regulatory 19 S subunit of the 26 S proteasome (5-7). It has been proposed that the latter and the signalosome have a common evolutionary origin but have diverged to assume different cellular functions.Moreover recent studies have shown an interaction of Fos/ Jun with SRC-1. The latter potentiates AP1 1 -mediated gene transactivation (8). SRC-1, also called p160 and ERAP160 (9 -12) is a member of the large family of nuclear receptor coactivators including SRC-2 (also called TIF2 and GRIP1) (13, 14), SRC-3 (also called TRAM1, ACTR, AIB1, RAC3, and p/CIP) (15-19), and ARA-70 (androgen receptor-associated protein) (20). These proteins form complexes with nuclear receptors and CBP or p300 (12,16,19,(21)(22)(23)(24). The latter recruit into transcription complexes histone acetylases such as p300/CBP-associated factor (16,25,26). Coactivators CBP and p300 also have intrinsic histone acetylase activity. Histo...
Cytokines and glucocorticoids (GCs) signaling pathways interfere with each other in the regulation of apoptosis and gene expression in the immune system. Interleukin-2 (IL-2), through the Janus kinase/signal transducers and activators of transcription (Jak/STAT) and mitogen-activated protein kinase (MAPK) pathways, activates STAT5 and activated protein-1 (AP-1) transcription factors, respectively, which are known to repress glucocorticoid receptor (GR) activity, at least in part, through protein-protein interactions. In this work, we have analyzed the mechanisms whereby IL-2 down-regulates the GC-induced transactivation of the mouse mammary tumor virus long terminal repeat (MMTV-LTR) in murine CTLL-2 T lymphocytes. Mutagenesis studies revealed that the MMTV-LTR STAT5 binding site (-923/-914) was not required for IL-2-mediated inhibition but identified both glucocorticoid response elements (GREs) and the -104/+1 region as critical elements for this negative response. The DNA binding activities of transcription factors required for GC-mediated activation of the MMTV-LTR promoter and that bind to the -104/+1 region (nuclear factor-1, Oct-1) were not affected by IL-2 treatment. Overexpression of wild-type STAT5B enhanced the effect of IL-2 on MMTV-LTR activity, and a dominant negative form of STAT5B (Y699F) abolished the IL-2-mediated MMTV-LTR inhibition, whereas AP-1 activation had no effect in this system. Direct interaction between liganded GR and STAT5 was observed in CTLL-2 cells in a STAT5 phosphorylation-independent manner. Overexpression of nuclear coactivators CBP (CREB-binding protein) or SRC-1a (steroid receptor coactivator 1a) did not blunt IL-2 inhibitory effects. We suggest that the STAT5-repressive activity on the GC-dependent transcription may involve direct interaction of STAT5 with GR, is dependent on the promoter context and STAT5 activation level, and occurs independently of coactivators levels in T cells.
We have used the gibbon ape leukemia cell line MLA-144 and its corticoid-sensitive subclone MLA-E7T to analyze the mechanisms whereby interleukin-2 (IL-2) can protect T cells against dexamethasone-induced apoptosis. MLA cells are characterized by the constitutive expression of intermediate affinity receptors for IL-2, together with IL-4 receptors. MLA-144 cells secrete IL-2 and are insensitive to dexamethasone, whereas MLA-E7T cells do not constitutively produce significant amounts of IL-2 and undergo apoptotic cell death in the presence of dexamethasone. Exogenous IL-2 was shown to protect MLA-E7T cells against the apoptotic effect of dexamethasone and to increase both the DNA binding and transactivating functions of activator protein-1 (AP-1). The functional relationship between AP-1 and glucocorticoid receptors transcriptional activities was further investigated using transient expression of reporter gene constructs whose transcriptions are regulated by promoters containing TPA-responsive elements or glucocorticoid-responsive elements. The data reported here demonstrate that in MLA-144 cells, IL-2 or PMA stimulation antagonizes the glucocorticoid receptor, whereas in MLA-E7T, synergistic effects are observed between dexamethasone and IL-2 or PMA for transactivation of MMTV-CAT. Taken together with the finding that IL-2 but not PMA protects MLA-E7T from dexamethasone-induced apoptosis, our results indicate that IL-2 does not induce such a protection by repressing the transcriptional activity of the glucocorticoid receptor.
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