Development of a Cell-Based Screen for Inhibitors of the FOG-1-NuRD Interaction

University Of Pennsylvania, Philadelphia PA

Investigators

Abstract

DESCRIPTION (provided by applicant): Sickle cell anemia is a serious disorder caused by a point mutation in the adult 2-globin gene. Patients with linked mutations that result in elevated levels of 3-globin, which is normally expressed only during fetal development, experience a more benign clinical course of the disease. This observation provided the initial impetus to better understand the mechanisms of 3-globin gene silencing with the goal to develop new treatments that would raise 3-globin levels. Based on recent findings obtained in our laboratory, we propose to develop a high-throughput screen to identify compounds that interfere with the function of a transcription factor complex that is involved in the silencing of 3-globin expression. The screen will be validated in 384- or 1536-well format in preparation for high-throughput screening (HTS) in the Molecular Libraries Probe Production Centers Network (MLPCN). The assay will be based on the repressive function of a transcription co-factor called FOG-1, which in turn binds to a co-repressor complex called NuRD (nucleosome remodeling and deacetylase). We defined a highly conserved module consisting of just 12 amino acids at the extreme N-terminus of FOG-1 that is required for NuRD binding. Disruption of just one amino acid within this motif leads to a marked reduction in NuRD binding and loss of repressor activity of FOG-1. This indicates that the contact surface between FOG-1 and NuRD is small. Therefore, we believe that it is possible to disrupt this interaction with small molecules. If successful, these compounds will be evaluated for their ability to reactivate 3-globin in an established transgenic mouse model and in human erythroid cells. Compounds that disrupt the FOG-1/NuRD interaction and raise the expression of 3-globin could provide a targeted treatment for sickle cell anemia and 2-thalassemia. Current treatment protocols include histone deacetylase inhibitors and DNA demethylating agents, which give rise to side effects resulting from very broad and rather unpredictable effects on cellular gene expression. Inhibitors of the FOG-1/NuRD interaction would be extremely useful chemical probes to dissect the transcriptional pathways employed by these nuclear factors and to further understanding of the mechanisms of hemoglobin switching. Moreover, since other important transcription repressors share this NuRD-binding module, chemicals that interfere with their function would be powerful tools to dissect transcriptional pathways during mammalian development. PUBLIC HEALTH RELEVANCE: Despite decades of intense studies, the mechanisms of hemoglobin switching are still unclear. Recent findings in our laboratory revealed a critical role for the FOG-1/NuRD transcription factor complex in the silencing of 3-globin expression. Compounds that target the FOG-1/NuRD interaction would be extremely useful tools to dissect the transcriptional pathways employed by these nuclear factors. Raising 3-globin levels by disruption of the FOG-1/NuRD complex may provide a novel, targeted treatment for sickle cell anemia and 2-thalassemia.