Description:

The Department of Biochemistry and Molecular Genetics Departmental Seminar Series presents:

Philip Iannaccone, MD, PhD
George M. Eisenberg Professor of Pediatrics, Northwestern University Feinberg School of Medicine

Director, Developmental Biology Program; Co-director, Human iPS and Stem Cell Core Facility; Director, Training Program, Stanley Manne Children's Research Institute, Ann & Robert H. Lurie Children’s Hospital of Chicago

In humans, a large cancer burden is associated with dysregulation of the Hedgehog/Patched/GLI (HH/PCTH/GLI) pathway. HH signaling is believed to be active in up to one-third of all human cancers. The canonical Hedgehog (including Sonic Hedgehog; SHH) signal is mediated by the GLI family of transcription factors. The SHH signal transduction pathway plays an important role in normal development and differentiation. GLI1 is one of three transcription factors that mediate SHH signal and is a human oncogene with gene targets that sustain proliferation, inhibit apoptosis, and promote angiogenesis as well as tumor cell migration. The tumor suppressor p53 normally competes with GLI1 for TAF9, which binds to transcription factors to promote assembly of transcriptional machinery. Excessive GLI1 inhibits p53’s tumor suppressor activity in part by competing TAF9 away from P53. GLI1 and GLI2 form a positive-feedback loop. We identified 6 GLI binding sites (GBS) in the large first intron of the human GLI1 gene that are highly conserved among mammals, and associated with histone H3K27Ac marks and DNase hypersensitivity clusters. Purified partial GLI1 and GLI2 bind all of the GBS. GLI1 multimerizes and binds the pioneer protein CBP. ChIP qPCR shows SHH signal dependent (Smoothened agonist) occupancy of the 1st intron by GLI2, and increased histone H3K4me3 occupancy in the 1st intron. Elimination of some of the GBS attenuates transcriptional activation. Eliminating all sites eliminates reporter gene activation. SHH stimulates occupancy near the sites by H3K27Ac, H3K4me3 and the histone bromodomain reader protein BRD4. BRD4 can be driven off the intron and TSS region with small inhibitory molecule I-BET151 and this results in a diminished H3K27Ac and H3K4Me3 occupancy. Further, GLI1 binds the histone variant H2A.Z. These results suggest that GLI1 and GLI2 could auto-regulate GLI1 through protein-protein interactions involving complexes of the transcription factors, histone variants, and reader proteins in the 1st intron of GLI1. The COSMIC project has already revealed a number of non-coding variants within the GLI1 1st intron that are associated with a variety of cancers. CRISPR/Cas9 editing of the GBS in human stem cells significantly reduces GLI1 target gene expression but also expression of GLI1 itself.