An artificial transcription factor that activates potent interferon-γ expression in human Jurkat T Cells
Interferon (IFN)-γ is a central regulator of cell-mediated immunity in human health and disease, but reduced expression of the target receptors impairs signaling activity and leads to immunotherapy resistance. Although intracellular expression of IFN-γ restores the signaling and downstream functions...
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2025-01-01
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| Series: | Frontiers in Molecular Medicine |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fmmed.2024.1492370/full |
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| author | Ashley King Davis Noblitt Olivia Sherron Olivia Sherron Clara Kjerfve Lydia Pless Nicholas L. Truex |
| author_facet | Ashley King Davis Noblitt Olivia Sherron Olivia Sherron Clara Kjerfve Lydia Pless Nicholas L. Truex |
| author_sort | Ashley King |
| collection | DOAJ |
| description | Interferon (IFN)-γ is a central regulator of cell-mediated immunity in human health and disease, but reduced expression of the target receptors impairs signaling activity and leads to immunotherapy resistance. Although intracellular expression of IFN-γ restores the signaling and downstream functions, we lack the tools to activate the IFNG gene instead of cell surface receptors. This paper introduces the design and characterization of an artificial transcription factor (ATF) protein that recognizes the IFNG gene with six zinc finger domains, which are dovetailed to a VP64 signaling domain that promotes gene transcription and translation. Biological studies with human Jurkat T cells reveal that the ATF amplifies IFNG gene transcription and translation, and also stimulates gene transcription for multiple class I and II HLA alleles and interferon-stimulated genes (ISGs). Biophysical characterization showed the recombinant ATF protein recognizes the human IFNG gene with nanomolar affinity (KD = 5.27 ± 0.3 nM), adopts a protein secondary structure associated with the ββα-fold of zinc finger domains, and is resistant to thermal denaturation. These studies demonstrate that transcriptional targeting of cytokine genes, rather than surface receptors, activates cytokine expression and shows significant potential for directing immune function. |
| format | Article |
| id | doaj-art-5ab0259a87194f46a945764a53f14dc6 |
| institution | Kabale University |
| issn | 2674-0095 |
| language | English |
| publishDate | 2025-01-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Molecular Medicine |
| spelling | doaj-art-5ab0259a87194f46a945764a53f14dc62025-01-08T06:12:04ZengFrontiers Media S.A.Frontiers in Molecular Medicine2674-00952025-01-01410.3389/fmmed.2024.14923701492370An artificial transcription factor that activates potent interferon-γ expression in human Jurkat T CellsAshley King0Davis Noblitt1Olivia Sherron2Olivia Sherron3Clara Kjerfve4Lydia Pless5Nicholas L. Truex6Department of Chemistry and Biochemistry, University of South Carolina, Columbia, SC, United StatesDepartment of Chemistry and Biochemistry, University of South Carolina, Columbia, SC, United StatesDepartment of Chemistry and Biochemistry, University of South Carolina, Columbia, SC, United StatesCollege of Engineering and Computing, University of South Carolina, Columbia, SC, United StatesDepartment of Chemistry and Biochemistry, University of South Carolina, Columbia, SC, United StatesDepartment of Chemistry and Biochemistry, University of South Carolina, Columbia, SC, United StatesDepartment of Chemistry and Biochemistry, University of South Carolina, Columbia, SC, United StatesInterferon (IFN)-γ is a central regulator of cell-mediated immunity in human health and disease, but reduced expression of the target receptors impairs signaling activity and leads to immunotherapy resistance. Although intracellular expression of IFN-γ restores the signaling and downstream functions, we lack the tools to activate the IFNG gene instead of cell surface receptors. This paper introduces the design and characterization of an artificial transcription factor (ATF) protein that recognizes the IFNG gene with six zinc finger domains, which are dovetailed to a VP64 signaling domain that promotes gene transcription and translation. Biological studies with human Jurkat T cells reveal that the ATF amplifies IFNG gene transcription and translation, and also stimulates gene transcription for multiple class I and II HLA alleles and interferon-stimulated genes (ISGs). Biophysical characterization showed the recombinant ATF protein recognizes the human IFNG gene with nanomolar affinity (KD = 5.27 ± 0.3 nM), adopts a protein secondary structure associated with the ββα-fold of zinc finger domains, and is resistant to thermal denaturation. These studies demonstrate that transcriptional targeting of cytokine genes, rather than surface receptors, activates cytokine expression and shows significant potential for directing immune function.https://www.frontiersin.org/articles/10.3389/fmmed.2024.1492370/fulltranscription factorinterferon-gammaenzyme-linked immunosorbent assay (ELISA)Jurkatimmunotherapyprotein design and engineering |
| spellingShingle | Ashley King Davis Noblitt Olivia Sherron Olivia Sherron Clara Kjerfve Lydia Pless Nicholas L. Truex An artificial transcription factor that activates potent interferon-γ expression in human Jurkat T Cells Frontiers in Molecular Medicine transcription factor interferon-gamma enzyme-linked immunosorbent assay (ELISA) Jurkat immunotherapy protein design and engineering |
| title | An artificial transcription factor that activates potent interferon-γ expression in human Jurkat T Cells |
| title_full | An artificial transcription factor that activates potent interferon-γ expression in human Jurkat T Cells |
| title_fullStr | An artificial transcription factor that activates potent interferon-γ expression in human Jurkat T Cells |
| title_full_unstemmed | An artificial transcription factor that activates potent interferon-γ expression in human Jurkat T Cells |
| title_short | An artificial transcription factor that activates potent interferon-γ expression in human Jurkat T Cells |
| title_sort | artificial transcription factor that activates potent interferon γ expression in human jurkat t cells |
| topic | transcription factor interferon-gamma enzyme-linked immunosorbent assay (ELISA) Jurkat immunotherapy protein design and engineering |
| url | https://www.frontiersin.org/articles/10.3389/fmmed.2024.1492370/full |
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