The nuclear receptor LRH-1 discriminates between ligands using distinct allosteric signaling circuits

Abstract

Nuclear receptors (NRs) are transcription factors that regulate essential biological processes in response to cognate ligands. An important part of NR function involves ligand-induced conformational changes that recruit coregulator proteins to the activation function surface (AFS), ~15 angstroms away from the ligand binding pocket. Ligands must communicate with the AFS to recruit appropriate coregulators and elicit different transcriptional outcomes, but this communication is poorly understood. These studies illuminate allosteric communication networks underlying activation of liver receptor homolog-1 (LRH-1), a NR that regulates development, metabolism, cancer progression and intestinal inflammation. Using >100 microseconds of all-atom molecular dynamics simulations involving 69 LRH-1 complexes, we identify distinct signaling circuits used by active and inactive ligands for AFS communication. Inactive ligands communicate via strong, coordinated motions along paths through the receptor to the AFS. Activating ligands disrupt the 'inactive' circuit by inducing connectivity elsewhere. Ligand-contacting residues in helix 7 help mediate the switch between circuits, suggesting new avenues for developing LRH-1-targeted therapeutics. We also elucidate aspects of coregulator signaling, showing that localized, destabilizing fluctuations are induced by inappropriate ligand-coregulator pairings. These studies have uncovered novel features of LRH-1 allostery, and the quantitative approach used to analyze many simulations provides a framework to study allosteric signaling in other receptors.

A novel heuristic of rigid docking scores positively correlates with full-length nuclear receptor LRH-1 regulation.

“…We found ∆∆G positively associated with the ability of our set of 58 hit compounds to regulate full-length LRH-1 in cellbased assays. Network analyses suggest the position of Helix 6 as a structural element that may contribute to the relationship between ∆∆G and ligand-mediated LRH-1 regulation in cells, consistent with analyses by independent groups [37,64,70,71,[74][75][76]. The data presented here suggest a role for Helix 6 in ligand-mediated regulation of LRH-1, similar to other studies which have applied comparative crystallography, MD simulations and network analyses [37,64,70,71,[74][75][76].…”

“…Network analyses suggest the position of Helix 6 as a structural element that may contribute to the relationship between ΔΔG and ligand-mediated LRH-1 regulation in cells, consistent with analyses by independent groups[37,64,70,71,74–76]. The data presented here suggest a role for Helix 6 in ligand-mediated regulation of LRH-1, similar to other studies which have applied comparative crystallography, MD simulations and network analyses[37,64,70,71,74–76]. The ΔΔG metric can help prioritize hit compounds for expensive secondary screening, which can hasten LRH-1 compound development.…”

“…Perhaps most importantly, convincing work from the Ortlund lab at Emory and now the Okafor lab at Penn State [74] have used extensive MD simulations, network analyses and direct comparisons of different ligand-bound structures of LRH-1 to independently suggest an important role for Helix 6 in ligand-regulated activation of LRH-1 [37,64,70,71,[74][75][76], linking the mobility of residues at the entrance to the ligand binding pocket near Helix 6 to synthetic small molecule-induced LRH-1 activity [76]. The volume of the ligand-binding pocket is smaller in structures bound to synthetic small molecules vs. phospholipids [64], consistent with the docking studies presented here, as well as hydrogen-deuterium exchange data [37,64,71,76], MD simulations [64,70,71,[74][75][76] and other network analyses [70,76]. Together, these studies support a role for Helix 6 is an important element in translating ligand binding events from the binding pocket to the transcriptional coregulator [70,71].…”

“…We then correlated those docking results retrospectively with published cell-based functional data from our previous study (439 total functional assay measurements) [1]. We found that docking to the full-length model of LRH-1 [73] predicts different binding positions for compounds active on full-length LRH-1, which were closer to the entrance of the ligand binding pocket and Helix 6, an important helix that helps mediate ligand activation of LRH-1 [37,64,70,71,[74][75][76]. The scores of compounds docked to full-length LRH-1 or the isolated LBD did not correlate with LRH-1 regulation in wet lab assays, however several compounds had large differences in docking scores (∆G) to the full-length vs. isolated LBD of LRH-1, an arbitrary metric we call ∆∆G.…”