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Updated: Jun 10, 2025
Identification of Transcription Factor Regulators using Medium-Throughput Screening of Arrayed Libraries and a Dual-Luciferase-Based Reporter
Published on: March 27, 2020
A High-Throughput Screen for Antiproliferative Peptides in Mammalian Cells Identifies Key Transcription Factor
Shane M Liila-Fogarty1,2, Grace E Boyum2,3, Claire L Schwabe1,2
1Department of Biomolecular Chemistry, University of Wisconsin-Madison, Madison, Wisconsin 53705, United States.
Abstract:
Transcription factors (TFs) are a promising therapeutic target for a multitude of diseases. TFs perform their cellular roles by participating in multiple specific protein-protein interactions. For example, homo- or heterodimerization of some TFs controls DNA binding, while interactions between TFs and components of basal transcriptional machinery or chromatin modifiers can also be critical. While, in theory, small molecules could be used to disrupt specific protein-protein interfaces required for TF function, in practice, it is difficult to identify small molecules with the necessary specificity and efficacy, likely due to the extensive protein-protein interfaces that often underlie TF function. However, in contrast to small molecules, peptides have the potential to provide both the specificity and efficacy required to disrupt such interfaces. Here, we identified ∼15 peptides that inhibit the proliferation of leukemia cells using a high-throughput pooled screen of a library of 80-mer protein regions (peptides) derived from human nuclear-localized proteins. The antiproliferative peptides were enriched for regions known to be involved in specific TF dimerization, including the basic leucine zipper (bZIP) domain family. One of these bZIP domains, JDP2;bZIP_1, from the TF JDP2, was the top antiproliferative peptide, reducing the proliferation of K562 cells by 2-fold. JDP2;bZIP_1 inhibited AP-1 transcriptional activity and phenocopied JDP2 overexpression, suggesting that the peptide affected proliferation through a native JDP2 mechanism. Unexpectedly, given the strong conservation of the bZIP domain, residues outside of the annotated dimerization domain were critical for the peptide's antiproliferative potency. The peptide-mediated antiproliferative effect initiated erythrocyte differentiation in K562 cells and increased G0/G1 cells across multiple cell line models. We also found that many of the antiproliferative peptides identified in this study, including JDP2;bZIP_1, did not require a nuclear localization signal to function, a potential benefit for delivering these peptides in therapeutic applications.
Insights
Researchers identified novel peptides that inhibit leukemia cell proliferation by targeting transcription factor dimerization. One peptide, JDP2;bZIP_1, effectively reduced cancer cell growth and promoted differentiation, offering a promising therapeutic strategy.
Area of Science:
- Molecular Biology
- Cancer Therapeutics
- Drug Discovery
Background:
- Transcription factors (TFs) are crucial for cellular function and implicated in various diseases.
- Targeting TF protein-protein interactions is a therapeutic strategy, but small molecules often lack specificity.
- Peptides offer potential for specific disruption of TF interfaces.
Purpose of the Study:
- To identify peptides that inhibit leukemia cell proliferation by targeting TF interactions.
- To characterize the mechanism of action of identified antiproliferative peptides.
- To explore the therapeutic potential of these peptides in cancer treatment.
Main Methods:
- High-throughput pooled screening of 80-mer peptides from human nuclear proteins.
- Assessing peptide antiproliferative activity against leukemia cells (K562).
- Investigating peptide effects on TF activity, cell differentiation, and cell cycle progression.
Main Results:
- Identified approximately 15 antiproliferative peptides, enriched for TF dimerization domains like bZIP.
- The peptide JDP2;bZIP_1 significantly inhibited K562 cell proliferation and AP-1 transcriptional activity.
- Peptide potency was linked to residues outside the conserved bZIP domain, and peptides did not require nuclear localization for function.
Conclusions:
- Peptides can effectively inhibit TF function and cancer cell proliferation.
- JDP2;bZIP_1 demonstrates therapeutic potential by targeting TF JDP2 mechanisms and inducing differentiation.
- The non-requirement for nuclear localization enhances the therapeutic applicability of these identified peptides.
