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Schnurri-3 (KRC) interacts with c-Jun to regulate the IL-2 gene in T cells.

Oukka M, Wein MN, Glimcher LH - J. Exp. Med. (2004)

Bottom Line: In the immune system, AP-1 activity is highest in T cells, suggesting that a subset of T cell-specific coactivator proteins exist to selectively potentiate AP-1 function.Here, we describe that the expression of Schnurri-3, also known as kappa recognition component (KRC), is induced upon T cell receptor signaling in T cells and functions to regulate the expression of the interleukin 2 (IL-2) gene.KRC physically associates with the c-Jun transcription factor and serves as a coactivator to augment AP-1-dependent IL-2 gene transcription.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology and Infectious Diseases, Harvard School of Public Health, Boston, MA 02115, USA.

ABSTRACT
The activator protein 1 (AP-1) transcription factor is a key participant in the control of T cell proliferation, cytokine production, and effector function. In the immune system, AP-1 activity is highest in T cells, suggesting that a subset of T cell-specific coactivator proteins exist to selectively potentiate AP-1 function. Here, we describe that the expression of Schnurri-3, also known as kappa recognition component (KRC), is induced upon T cell receptor signaling in T cells and functions to regulate the expression of the interleukin 2 (IL-2) gene. Overexpression of KRC in transformed and primary T cells leads to increased IL-2 production, whereas dominant-negative KRC, or loss of KRC protein in KRC- mice, results in diminished IL-2 production. KRC physically associates with the c-Jun transcription factor and serves as a coactivator to augment AP-1-dependent IL-2 gene transcription.

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KRC overexpression increases the transcription of the IL-2 gene. (A) Stably transfected Jurkat T cell clones with vector (vec) or KRC (JURKAT-KRC) were stimulated with 50 ng/ml PMA plus 2 μM ionomycin for 6 h. IL-2 mRNA abundance was determined by RT-PCR with tubulin as an internal control. (B) Jurkat cells were transiently transfected with an IL-2–luciferase reporter (IL-2-LUC) along with Vector, KRC, or KRCtr (amino acids 204–1,055) and, in all cases, a CMV–β-Gal reporter as an internal control (Materials and Methods). 24 h later, cells were stimulated with PMA plus ionomycin for 6 h (top) or Raji cells loaded with SEE for 8 h (bottom). Luciferase activity was determined and normalized for β-galactosidase activity. (C) Jurkat cells were transiently transfected with NFAT/AP-1–, NFAT-, or AP-1–Luciferase reporters and treated as aforemtioned.
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fig3: KRC overexpression increases the transcription of the IL-2 gene. (A) Stably transfected Jurkat T cell clones with vector (vec) or KRC (JURKAT-KRC) were stimulated with 50 ng/ml PMA plus 2 μM ionomycin for 6 h. IL-2 mRNA abundance was determined by RT-PCR with tubulin as an internal control. (B) Jurkat cells were transiently transfected with an IL-2–luciferase reporter (IL-2-LUC) along with Vector, KRC, or KRCtr (amino acids 204–1,055) and, in all cases, a CMV–β-Gal reporter as an internal control (Materials and Methods). 24 h later, cells were stimulated with PMA plus ionomycin for 6 h (top) or Raji cells loaded with SEE for 8 h (bottom). Luciferase activity was determined and normalized for β-galactosidase activity. (C) Jurkat cells were transiently transfected with NFAT/AP-1–, NFAT-, or AP-1–Luciferase reporters and treated as aforemtioned.

Mentions: The production of IL-2 by T cells is regulated at multiple levels, including transcription, mRNA stability, and rate of protein secretion (24, 25). To define at which stages KRC acts, we first measured levels of IL-2 mRNA transcripts by semi-quantitative RT-PCR in Jurkat T cells stably transfected with full-length KRC. As seen in Fig. 3 A, Jurkat clones overexpressing KRC displayed higher levels of IL-2 transcripts when activated than Jurkat clones transfected with vector control. Next, we tested whether KRC was able to directly transactivate a 1.5-kb IL-2 promoter-luciferase reporter in Jurkat cells. Provision of KRC resulted in an ∼10-fold induction of luciferase activity in Jurkat cells treated with PMA plus ionomycin (Fig. 3 B, top). Just as KRC overexpression alone did not lead to spontaneous production of endogenous IL-2, no transactivation by KRC was observed in the absence of PMA/ionomycin in these luciferase reporter assays. To provide a more physiologic signal to activate Jurkat cells, we used a model system in which Raji B lymphoma cells act as antigen-presenting cells to present SEE to Jurkat. As shown in Fig. 3 B (bottom), provision of KRC substantially increased (∼10-fold) IL-2 promoter activity in this system. Interestingly, KRC had no effect on IL-2 promoter activity in the absence of Jurkat activation either by PMA/ionomycin or by antigen/APC. These data further suggest that KRC expression alone is not sufficient to induce IL-2 mRNA expression; instead, KRC's ability to enhance IL-2 production relies on endogenous factors found only in activated T cells.


Schnurri-3 (KRC) interacts with c-Jun to regulate the IL-2 gene in T cells.

Oukka M, Wein MN, Glimcher LH - J. Exp. Med. (2004)

KRC overexpression increases the transcription of the IL-2 gene. (A) Stably transfected Jurkat T cell clones with vector (vec) or KRC (JURKAT-KRC) were stimulated with 50 ng/ml PMA plus 2 μM ionomycin for 6 h. IL-2 mRNA abundance was determined by RT-PCR with tubulin as an internal control. (B) Jurkat cells were transiently transfected with an IL-2–luciferase reporter (IL-2-LUC) along with Vector, KRC, or KRCtr (amino acids 204–1,055) and, in all cases, a CMV–β-Gal reporter as an internal control (Materials and Methods). 24 h later, cells were stimulated with PMA plus ionomycin for 6 h (top) or Raji cells loaded with SEE for 8 h (bottom). Luciferase activity was determined and normalized for β-galactosidase activity. (C) Jurkat cells were transiently transfected with NFAT/AP-1–, NFAT-, or AP-1–Luciferase reporters and treated as aforemtioned.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC1887724&req=5

fig3: KRC overexpression increases the transcription of the IL-2 gene. (A) Stably transfected Jurkat T cell clones with vector (vec) or KRC (JURKAT-KRC) were stimulated with 50 ng/ml PMA plus 2 μM ionomycin for 6 h. IL-2 mRNA abundance was determined by RT-PCR with tubulin as an internal control. (B) Jurkat cells were transiently transfected with an IL-2–luciferase reporter (IL-2-LUC) along with Vector, KRC, or KRCtr (amino acids 204–1,055) and, in all cases, a CMV–β-Gal reporter as an internal control (Materials and Methods). 24 h later, cells were stimulated with PMA plus ionomycin for 6 h (top) or Raji cells loaded with SEE for 8 h (bottom). Luciferase activity was determined and normalized for β-galactosidase activity. (C) Jurkat cells were transiently transfected with NFAT/AP-1–, NFAT-, or AP-1–Luciferase reporters and treated as aforemtioned.
Mentions: The production of IL-2 by T cells is regulated at multiple levels, including transcription, mRNA stability, and rate of protein secretion (24, 25). To define at which stages KRC acts, we first measured levels of IL-2 mRNA transcripts by semi-quantitative RT-PCR in Jurkat T cells stably transfected with full-length KRC. As seen in Fig. 3 A, Jurkat clones overexpressing KRC displayed higher levels of IL-2 transcripts when activated than Jurkat clones transfected with vector control. Next, we tested whether KRC was able to directly transactivate a 1.5-kb IL-2 promoter-luciferase reporter in Jurkat cells. Provision of KRC resulted in an ∼10-fold induction of luciferase activity in Jurkat cells treated with PMA plus ionomycin (Fig. 3 B, top). Just as KRC overexpression alone did not lead to spontaneous production of endogenous IL-2, no transactivation by KRC was observed in the absence of PMA/ionomycin in these luciferase reporter assays. To provide a more physiologic signal to activate Jurkat cells, we used a model system in which Raji B lymphoma cells act as antigen-presenting cells to present SEE to Jurkat. As shown in Fig. 3 B (bottom), provision of KRC substantially increased (∼10-fold) IL-2 promoter activity in this system. Interestingly, KRC had no effect on IL-2 promoter activity in the absence of Jurkat activation either by PMA/ionomycin or by antigen/APC. These data further suggest that KRC expression alone is not sufficient to induce IL-2 mRNA expression; instead, KRC's ability to enhance IL-2 production relies on endogenous factors found only in activated T cells.

Bottom Line: In the immune system, AP-1 activity is highest in T cells, suggesting that a subset of T cell-specific coactivator proteins exist to selectively potentiate AP-1 function.Here, we describe that the expression of Schnurri-3, also known as kappa recognition component (KRC), is induced upon T cell receptor signaling in T cells and functions to regulate the expression of the interleukin 2 (IL-2) gene.KRC physically associates with the c-Jun transcription factor and serves as a coactivator to augment AP-1-dependent IL-2 gene transcription.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology and Infectious Diseases, Harvard School of Public Health, Boston, MA 02115, USA.

ABSTRACT
The activator protein 1 (AP-1) transcription factor is a key participant in the control of T cell proliferation, cytokine production, and effector function. In the immune system, AP-1 activity is highest in T cells, suggesting that a subset of T cell-specific coactivator proteins exist to selectively potentiate AP-1 function. Here, we describe that the expression of Schnurri-3, also known as kappa recognition component (KRC), is induced upon T cell receptor signaling in T cells and functions to regulate the expression of the interleukin 2 (IL-2) gene. Overexpression of KRC in transformed and primary T cells leads to increased IL-2 production, whereas dominant-negative KRC, or loss of KRC protein in KRC- mice, results in diminished IL-2 production. KRC physically associates with the c-Jun transcription factor and serves as a coactivator to augment AP-1-dependent IL-2 gene transcription.

Show MeSH
Related in: MedlinePlus