LRH-1 (NR5A2) is a monomeric nuclear receptor involved in many aspects of liver physiology, including bile acid, cholesterol and glucose homeostasis. LRH-1 activation has beneficial effects on liver metabolism in pre-clinical mouse models. As nuclear receptors like LRH-1 have a very druggable ligand-binding pocket, LRH-1 has been targeted by many drug development efforts with great recent progress, however an LRH-1 agonist is still not available in the clinic. Like most other nuclear receptors, LRH-1 is composed of a DNA-binding domain and a ligand-binding domain, which are connected by a large unstructured Hinge domain. Classic nuclear receptor drug design has focused on the isolated ligand-binding domain, as the regulatory mechanism of this isolated domain is very well understood at the molecular level: binding of a hydrophobic small molecule allosterically alters ligand-binding domain recruitment of a transcriptional coregulator, which regulates nuclear receptor function. However, several lines of evidence suggest inter-domain communication exists between LRH-1 domains, regulating function. Understanding the structural biology behind this inter-domain communication might help LRH-1 drug design efforts, however technical challenges in applying crystallography or cryo-EM has prevented progress, despite great effort from several groups. We used an integrated structural approach to develop a low-resolution, but high confidence model of the intact, full-length LRH-1, using exclusively solution-based biophysical analyses and computational modeling (HDX, SAXS, chemical crosslinking, artificial amino acid benzophenone cross linking, Cys-Cys interdomain crosslinking, Rosetta and MD simulations). The model explains human patient mutations and structure-based mutations predicted to reside in the interface between the domains, which we show alter full length LRH-1 structure and function. Here, we propose to take advantage of this solution-based approach to address several long-standing questions in the field:
Aim 1 determines how various ligands change full length LRH-1 interdomain communication.
Aim 2 resolves how the SUMO module post-translational modification alters LRH-1 interdomain communication.
Aim 3 identifies the genes effected by structure-based LRH-1 mutations in mouse liver and primary hepatocytes. Our current understanding of how LRH-1 structure is regulated is limited to studies of the individual domains. Without understanding how full-length LRH-1 is regulated, we cannot know if our current drug design efforts are taking full advantage of the entire therapeutic capacity of LRH-1.
Detailed structural biology describes the isolated domains of the nuclear receptor Liver Receptor Homolog-1 (LRH-1, NR5A2) suggesting how LRH-1 can be targeted in NAFLD, NASH and type 2 diabetes, however very little structural biology describes full length LRH-1. We have developed an integrated structural approach to analyze full length LRH-1, which produced a low resolution, but high confidence model of full length LRH-1 structure in solution, which feasibly permits us to ask how full length LRH-1 structure is regulated. Thus this proposal helps ongoing drug development efforts by revealing how LRH-1 domain architecture is regulated to influence LRH-1 function, providing new structure-function relationships that make agonist development more likely to succeed.
