Interaction between the trout mineralocorticoid and glucocorticoid receptors in vitro

Abstract

The salmonid corticosteroid receptors (CRs), glucocorticoid receptors 1 and 2 (GR1 and GR2) and the mineralocorticoid receptor (MR) share a high degree of homology with regard to structure, ligand-and DNA response element-binding, and cellular co-localization. Typically, these nuclear hormone receptors homodimerize to confer transcriptional activation of target genes, but a few studies using mammalian receptors suggest some degree of heterodimerization. We observed that the trout MR confers a several fold lower transcriptional activity compared to the trout GRs. This made us question the functional relevance of the MR when this receptor is located in the same cells as the GRs and activated by cortisol. A series of co-transfection experiments using different glucocorticoid response elements (GREs) containing promoter-reporter constructs were carried out to investigate any possible interaction between the piscine CRs. Co-transfection of the GRs with the MR significantly reduced the total transcriptional activity even at low MR levels, suggesting interaction between these receptors. Co-transfection of GR1 or GR2 with the MR did not affect the subcellular localization of the GRs, and the MR-mediated inhibition seemed to be independent of specific activation or inhibition of the MR. Site-directed mutagenesis of the DNA-binding domain and dimerization interface of the MR showed that the inhibition was dependent on DNA binding but not necessarily on dimerization ability. Thus, we suggest that the interaction between MR and the GRs may regulate the cortisol response in cell types where the receptors co-localize and propose a dominant-negative role for the MR in cortisolmediated transcriptional activity.

30 YEARS OF THE MINERALOCORTICOID RECEPTOR: Evolution of the mineralocorticoid receptor: sequence, structure and function

“…Cortisol-activated MR-GR heterodimers bind to glucocorticoid response elements, regulating glucocorticoid target genes (Mifsud & Reul 2016). Recently, heterodimers between trout MR and GR were studied in detail, and the MR in the presence of either cortisol or DOC was found to be a dominant negative repressor of trout GR (Kiilerich et al 2015). Thus, the actions of the MR and GR in cells that coexpress both receptors are complex and can be influenced by steroids that bind both receptors or are selective for each receptor.…”

Transcriptional Activation of Elephant Shark Mineralocorticoid Receptor by Corticosteroids, Progesterone and Spironolactone

“…The use of Hek293 cells and an assay temperature of 37 C does not replicate the physiological environment of Elephant sharks. Nevertheless, studies with Hek293 cells and other mammalian cell lines have proven useful for other studies of transcriptional activation by corticosteroids of skate (61) and teleost fish (36,44,75,76) MRs.…”

The evolution, structure and function of the ray finned fish (Actinopterygii) glucocorticoid receptors

“…Interestingly, a recent study by Kiilerich et al (2015) 320 identified that the rainbow trout MR, which is also activated by cortisol and in the absence of 321 defined teleost mineralocorticoid or function for MR (Takahashi and Sakamoto, 2013) could 322 be described as a third teleost GR, is a repressor of both rtGR1 and rtGR2 transcriptional 323 activity (Figure 3). Similar observations have been made with the mammalian GR and MR 324 and the repressor activity is due to GR/MR heterodimerisation that disrupts the self-325 synergistic interactions between the N-terminal of 2 GRs when bound as a homodimer (Liu 326 et al 1995).…”

“…To date, the zebrafish are the only fish species 408 known to have "human-like" GRβ, but GR activity repression may occurs via a different route 409 in other fish species, with Kiilerich et al (2015) showing that in rainbow trout the MR, which 410 also binds cortisol and can activate GREs in vitro (Sturm et al 2005), can act to repress GR1 411…”