Regulation of transcription factors by XAP-2, a member of the Hsp90 chaperone complex

Abstract: p>The AhR belongs to the bHLH/PAS family of transcription factors, which mediates biological effects of environmental pollutants such as 2,3,7,8-tetrachlorodibenzo-rho-dioxin (TCDD). In the absence of ligand the AhR is located in the cytoplasmatic compartment of the cell associated with the molecular chaperone Hsp90, p23 and a member of the immunophilin family of proteins, XAP-2. Upon ligand binding the receptor translocates to the nuclear compartment of the cell where it dimerizes with its partner factor ARNT. This dimerization event induces the release of the Hsp90 complex and subsequent binding of the AhR/ARNT complex to promoter regions of TCDD regulated genes such as Cyp1A1 and Cyp1A2. Following transcriptional activation AhR is exported out of the nucleus by the export protein CRM1. CRM1 binds to nuclear export sequences (NES) within transcription factors and in turn mediates export of the AhR to the cytoplasm. In this study we show that nuclear export of the AhR is mediated by two different NES depending on ligand status. In paper I we show that in the absence of ligand, export of the AhR is mediated by a NES located in the PAS A domain of the AhR. In contrast, nuclear export of the ligand activated AhR is mediated by a NES located in the bHLH domain of the receptor. In paper II we show that cytoplasmatic localization of the AhR is mediated by two non-overlapping mechanisms. One is mediated through CRM1 and the second mechanism to maintain the AhR in the cytoplasm is dependent on the Hsp90 associated co-chaperone XAP-2. Here we show that XAP-2 mediates anchoring of the non-activated AhR to actin filaments in the cytoplasm. XAP-2 is a 37kDa Hsp90 associated protein that belongs to the immunophilin family of proteins such as FKBP51 and 52. It is mainly located in the cytoplasmatic compartment of the cell where it protects the AhR against proteosomal degradation by inhibit ubiquitinylation of the AhR in the cytosol. XAP-2 has previously been identified as an Hsp90 associated protein that specifically interacts with the AhR. This notion, however have been challenged upon findings that XAP-2 interacts with other proteins such as PPARalpha. In this study we show that XAP-2 interacts with both ERalpha and TRbeta1. Interestingly, this interaction is isoform specific. Furthermore, in paper III we show that XAP-2 regulates the transcription of hypothalamic thyrotropin releasing hormone (TRH) in vivo demonstrating an important regulatory and physiological role for an immunophilin like factor in vivo. In paper IV we observe that XAP-2 exerts a repressive effect on ERalpha but not ERbeta transcriptional activation. In addition, this effect is mediated by the co-activator TIF-2. Taken together our results show that XAP-2 function as a general co-factor that modulates numerous cellular regulatory pathways including the AhR, TR and ER. These studies provide new perspectives in the understanding of the regulation of transcription factors by members of the Hsp90 complex.