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Barbara Graves

Professor and Chair of Oncological Sciences

B.A. Rice University

Ph.D. University of Washington

Research

References

barbara.graves@utah.edu

Barbara Graves' Lab Page

Barbara Graves' PubMed Literature Search

Research

The Graves Laboratory focuses on mechanistic questions in the area of transcriptional regulation. We use biochemical, biophysical and molecular biology approaches to investigate how sequence-specific DNA binding transcription factors regulate gene expression in normal and diseased states. Current projects explore signaling pathways, DNA binding properties, and protein-protein interactions within the ETS family of regulatory transcription factors.

Regulation of gene expression requires the targeting of regulatory transcription factors to their site of action in the genome. Molecular complementarity between a regulatory protein and its DNA binding site provides the primary targeting mechanism. A constellation of electrostatic and hydrophobic interactions between matching surfaces of the DNA helix and the protein establish high-affinity and sequence-specific binding. The effectiveness of this macromolecular matchmaking is challenged by the complexities of eukaryotes. There are hundreds of regulatory transcription factors that function by binding DNA sequences within promoter regions. Almost all of these proteins are encoded by multi-gene families. Members of a family display the same structural fold for binding DNA and recognize similar DNA sequences. My laboratory investigates the regulatory pathways that provide specificity for transcription factors that belong to multi-gene families.

Our investigations are currently focused on the ETS gene family that dramatically illustrates the specificity problem. ETS genes are present in all metazoan phyla with 27 homologs in the human genome. Most cell types express at least 16 of the 27 ETS genes. The ETS domain, a highly conserved 85-amino acid region, defines the family and directs DNA binding to the core recognition sequence 5’-GGAA/T-3’. With such a high degree of conservation, we ask how is specificity programmed into the family? We are performing genome-wide searches for transcriptional targets of ETS proteins to determine the rules for specificity using the latest technology for high-through put sequencing. Unbiased searches for Ets-1 targets have discovered unexpected dual occupancy with other ETS proteins at some targets as well as specific occupancy at other targets. Bioinformatics approaches are identifying the sequence motifs that distinguish these dual roles. The specific targets are characterized by a binding site for a partner DNA binding protein RUNX1 that adds sequence preferences, and thus, additional sequence specificity. This combination of factors also directs the recruitment of the co-activator CBP/p300.

We also use the ETS family to understand how dysregulation of gene expression occurs during human cancers. Nuclear oncogenes are most often transcription factors that cause inappropriate gene expression. The ets family illustrates this oncogenic potential. Members of the family are targets of Ras-dependent signaling, a growth control pathway frequently mutated to be superactivated in human cancers. We have characterized how Ras-dependent signaling leads to phosphorylation of Ets-1. The effect of signaling is enhancement of transcriptional activity. We discovered that the co-activator CBP/p300 binding is enhanced by phosphorylation, thus Ras/MAPK signaling to Ets-1 functions at the level of co-activator recruitment. We are currently studying this interaction at a structural level to understand how the commonly used co-activators, such as CBP/p300, are specifically recruited to Ets-1 target genes. We have discovered that phosphorylation affects both the structure of Ets-1 and adds negative charges to enhance the electrostatic forces that drive the interaction. We plan to screen for small molecules that can disrupt these molecular interactions in an effort to identify novel drug targets within the Ras/MAPK pathway.

References