Cellular transcription programs require that many proteins of different function assemble on chromatin in response to signals that originate outside the nucleus. The largest family of proteins initiating these transcriptional processes is that of the nuclear receptors. The nuclear receptors are multidomain proteins, and there are no atomic resolution images of their complete structures. Our proposal focuses on the nuclear receptor LRH-1 (Liver Receptor Homologue 1, termed also Pancreas Receptor Homologue 1), a critical transcription factor found in liver, intestines and pancreas. Our ultimate goal is to image the full length LRH-1 receptor with partner co-regulatory proteins when bound to DNA. We will use an advanced methodology to prepare and assemble the multiple domains of LRH-1 with partner proteins and specific DNA fragments so that the transcriptional assembly would be ordered well enough for analysis by X-ray crystallography. We plan to employ a stabilizing transcriptional partner protein, beta-catenin, that will permit biochemical preparation of previously intractable molecules and mediate their interactions in the functional complexes. We will use a systematic approach to identify stable assemblies of LRH-1 with its characterized transcriptional regulators. Specific DNA fragments representing known response elements of LRH-1 will be included in evaluation of our assemblies. Methods that we develop for imaging the regulatory complexes built by LRH-1 will be directly applicable to other nuclear receptors and their transcriptional assemblies. Our goal is to take a major step in learning how to determine structures for functional nuclear receptors and learn atomic level details about the mechanisms of their assembly and regulation of transcription. One third of current pharmaceuticals target regulatory sites of the nuclear receptors because of their primary roles in RNA transcription in all biological processes. Novel structural information about the transcriptional machines built by nuclear receptors on DNA would vastly expand our understanding of transcription and its regulation as well as physiological and pathophysiological pathways controlled by these transcriptional processes. Our proposal focuses on the transcriptional assemblies built by nuclear receptor LRH-1 and its co-regulator beta-catenin. We chose LRH-1 as a primary target because of the receptor's critical roles in human developmental, metabolic and numerous pathophysiological processes. As we describe in our proposal, deregulation of LRH-1 in pancreas is linked to maturity-onset diabetes. Functional LRH-1/beta-catenin deficiencies lead to inflammatory bowel disease, Crohn's disease and ulcerative colitis. In contrast, the aberrant, up-regulated activity of the LRH-1/ beta-catenin partnership is implicated in triggering and proliferation of cancerous tumors, including gastrointestinal cancers. Therefore, we expect that the atomic resolution architecture of the functional LRH-1/beta-catenin assembly will propel drug discovery process and enable the development of novel potent therapeutics for targeted intervention therapies.
We are studying nuclear receptors - protein molecules that initiate changes in gene transcription and therefore play primary roles in all biological processes. We concentrate our efforts on challenging goal of imaging the nuclear receptor LRH-1, a critical transcription factor found in liver, intestines and pancreas, with its transcriptional regulatory partners when bound on DNA. We expect that imaging of functional regulatory assemblies formed by this receptor will not only vastly expand our understanding of transcription and its regulation but also enable the development of potent therapeutics for targeted treatments of numerous diseases, including chronic intestinal inflammation and gastrointestinal and pancreatic cancers.
| Yumoto, Fumiaki; Nguyen, Phuong; Sablin, Elena P et al. (2012) Structural basis of coactivation of liver receptor homolog-1 by ?-catenin. Proc Natl Acad Sci U S A 109:143-8 |
