Hematopoiesis, the differentiation of distinct lineages of blood cells from a common progenitor, is intricately controlled at the transcriptional level. Detailed knowledge of the transcription factors and their molecular interactions is essential to a complete understanding of normal hematopoiesis as well as an array of disorders that arise from de-regulated transcriptional control. In addition, transcriptional regulation in hematopoiesis is a paradigm for reprogramming pluripotent stem cells and is therefore a key area of inquiry in bioengineering as well. Currently, hematopoietic transcription factors are modulated indirectly through their expression levels by genetic approaches, such as transgenes, engineered knock-out/down animal models and RNA interference. These techniques, which are slow in onset, persistent in duration, and global in effect, are ill-suited to dissect transcriptional pathways an probe transient interactions that typify transcriptional control. Direct chemical control of transcriptional activity i.e. at the protein/DNA level, is currently unavailable for the vast majorty of hematopoietic regulators. Additionally the strong structural conservation among members of transcription factor families confounds simple screening of chemical libraries against these transcriptional factors. Here, we explore a physicochemical strategy to isolate activators of structurally homologous members of the ETS-family transcription factors PU.1 and Ets-1, which play distinct roles in hematopoietic cell-fate determination. Specifically, we will screen a phage display library against cognate PU.1/DNA and Ets-1/DNA complexes under osmotically variable conditions (alternating conditions of normal and elevated osmolality) to isolate peptides that selectively stabilize the binding of PU.1 or Ets-1 to their cognate binding sites. Our proposal is based on our recent studies showing that cognate site binding by PU.1 is exquisitely sensitive to osmotic stress, but Ets-1 is insensitive, even though the DNA binding domains of the two proteins are structurally superimposable. After further optimizing sequence candidates using combinatorial libraries, we will determine the ability of lead candidates to localize in the nucleiof hematopoietic progenitor cells and specifically activate PU.1 or Ets-1 target genes. Given the emerging interest in PU.1 and Ets-1 in a broad range of experimental models of hematopoiesis, the peptides generated from this research are expected to become useful reagents for interrogating PU.1 and Ets-1 target genes as well as templates for targeted diagnostics and therapeutics in hematologic, rheumatologic, and infectious diseases.

Public Health Relevance

This proposal is aimed at developing new compounds that act directly on and activate cellular regulators involved in the development of blood cells from their precursors. These compounds are designed to overcome limitations of existing genetic methods and enable previously inaccessible experimental studies into the function and interactions of these essential regulators. In addition, we anticipate these novel compounds to serve as templates for new diagnostics and therapeutics for hematologic, rheumatologic, and infectious diseases in which these regulators have been recently implicated.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Exploratory/Developmental Grants (R21)
Project #
1R21HL129063-01
Application #
8947574
Study Section
Macromolecular Structure and Function B Study Section (MSFB)
Program Officer
Thomas, John
Project Start
2015-09-15
Project End
2017-06-30
Budget Start
2015-09-15
Budget End
2016-06-30
Support Year
1
Fiscal Year
2015
Total Cost
$201,000
Indirect Cost
$76,000
Name
Georgia State University
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
837322494
City
Atlanta
State
GA
Country
United States
Zip Code
30302
Albrecht, Amanda V; Kim, Hye Mi; Poon, Gregory M K (2018) Mapping interfacial hydration in ETS-family transcription factor complexes with DNA: a chimeric approach. Nucleic Acids Res 46:10577-10588
Mao, Peng; Brown, Alexander J; Esaki, Shingo et al. (2018) ETS transcription factors induce a unique UV damage signature that drives recurrent mutagenesis in melanoma. Nat Commun 9:2626
Antony-Debré, Iléana; Paul, Ananya; Leite, Joana et al. (2017) Pharmacological inhibition of the transcription factor PU.1 in leukemia. J Clin Invest 127:4297-4313
Poon, Gregory M K; Kim, Hye Mi (2017) Signatures of DNA target selectivity by ETS transcription factors. Transcription 8:193-203
Vo, Tam; Wang, Shuo; Poon, Gregory M K et al. (2017) Electrostatic control of DNA intersegmental translocation by the ETS transcription factor ETV6. J Biol Chem 292:13187-13196
Esaki, Shingo; Evich, Marina G; Erlitzki, Noa et al. (2017) Multiple DNA-binding modes for the ETS family transcription factor PU.1. J Biol Chem 292:16044-16054
Erlitzki, Noa; Huang, Kenneth; Xhani, Suela et al. (2017) Investigation of the electrostatic and hydration properties of DNA minor groove-binding by a heterocyclic diamidine by osmotic pressure. Biophys Chem 231:95-104
Xhani, Suela; Esaki, Shingo; Huang, Kenneth et al. (2017) Distinct Roles for Interfacial Hydration in Site-Specific DNA Recognition by ETS-Family Transcription Factors. J Phys Chem B 121:2748-2758
Stephens, Dominique C; Poon, Gregory M K (2016) Differential sensitivity to methylated DNA by ETS-family transcription factors is intrinsically encoded in their DNA-binding domains. Nucleic Acids Res 44:8671-8681
Stephens, Dominique C; Kim, Hye Mi; Kumar, Arvind et al. (2016) Pharmacologic efficacy of PU.1 inhibition by heterocyclic dications: a mechanistic analysis. Nucleic Acids Res 44:4005-13