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The Ikaros transcription factor regulates responsiveness to IL-12 and expression of IL-2 receptor alpha in mature, activated CD8 T cells.

Clambey ET, Collins B, Young MH, Eberlein J, David A, Kappler JW, Marrack P - PLoS ONE (2013)

Bottom Line: While DN Ikaros-transduced cells had higher expression of the IL-2 receptor alpha chain, DN Ikaros-transduced cells achieved their competitive advantage through an IL-2 independent mechanism.Finally, the competitive advantage of DN Ikaros-transduced cells was manifested in vivo, following adoptive transfer of transduced cells.These data identify the Ikaros family of transcription factors as regulators of cytokine responsiveness in activated CD8 T cells, and suggest a role for this family in influencing effector and memory CD8 T cell differentiation.

View Article: PubMed Central - PubMed

Affiliation: Integrated Department of Immunology, University of Colorado Anschutz Medical Campus, Aurora, Colorado, USA. eric.clambey@ucdenver.edu

ABSTRACT
The Ikaros family of transcription factors is critical for normal T cell development while limiting malignant transformation. Mature CD8 T cells express multiple Ikaros family members, yet little is known about their function in this context. To test the functions of this gene family, we used retroviral transduction to express a naturally occurring, dominant negative (DN) isoform of Ikaros in activated CD8 T cells. Notably, expression of DN Ikaros profoundly enhanced the competitive advantage of activated CD8 T cells cultured in IL-12, such that by 6 days of culture, DN Ikaros-transduced cells were 100-fold more abundant than control cells. Expression of a DN isoform of Helios, a related Ikaros-family transcription factor, conferred a similar advantage to transduced cells in IL-12. While DN Ikaros-transduced cells had higher expression of the IL-2 receptor alpha chain, DN Ikaros-transduced cells achieved their competitive advantage through an IL-2 independent mechanism. Finally, the competitive advantage of DN Ikaros-transduced cells was manifested in vivo, following adoptive transfer of transduced cells. These data identify the Ikaros family of transcription factors as regulators of cytokine responsiveness in activated CD8 T cells, and suggest a role for this family in influencing effector and memory CD8 T cell differentiation.

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CD8 T cells expressing dominant negative Ikaros have a modest advantage in vivo.(A) Relative abundance of retrovirally transduced cells within a pool of adoptively transferred CD8 T cells containing both transduced and non-transduced cells. Adoptively transferred cells were identified as live, CD8+ MHC class II negative cells that expressed CD45.1, with retrovirally transduced cells further expressing Thy1.1, with abundance measured within the peripheral blood of C57BL/6 recipient mice (which do not express CD45.1 or Thy1.1). Cells were transduced with a control retrovirus (white bar) or a DN Ikaros expressing retrovirus (black bar). To standardize for slightly different starting frequencies, values are standardized relative to day 1 input values for either control or DN Ikaros transduction efficiencies with data showing mean ± SEM with 3–10 mice for control transduced and 6–10 mice for DN Ikaros transduced cells. Mice were bled on day 1, 7, and 39 post-transfer. Statistically significant differences (p<0.05) are indicated, calculated by unpaired t test. (B) Fold change in the abundance of retrovirally transduced cells in C57BL/6J recipient mice from panel A (either control or DN Ikaros transduced) in the blood (defined as the percentage of live, CD8+ MHC class II negative cells that were CD45.1+ Thy1.1+) following injection of either vehicle alone or recombinant IL-12 (1 µg) at day 162 and 165 post-adoptive transfer, with abundance post-treatment measured at day 168 post-transfer. For these studies, at day 1 after adoptive transfer, the frequency of adoptively transferred, retrovirally transduced cells (defined as CD45.1+ Thy1.1+) among CD8 T cells in the peripheral blood was 1.21±0.07% for control and 1.70±0.09% for DN Ikaros transduced cells (mean ± SEM). Among adoptively transferred cells (CD45.1+), the percentage (mean ± SEM) of transduced cells was 42.1±0.7% for control and 49.6±0.5% for DN Ikaros.
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pone-0057435-g006: CD8 T cells expressing dominant negative Ikaros have a modest advantage in vivo.(A) Relative abundance of retrovirally transduced cells within a pool of adoptively transferred CD8 T cells containing both transduced and non-transduced cells. Adoptively transferred cells were identified as live, CD8+ MHC class II negative cells that expressed CD45.1, with retrovirally transduced cells further expressing Thy1.1, with abundance measured within the peripheral blood of C57BL/6 recipient mice (which do not express CD45.1 or Thy1.1). Cells were transduced with a control retrovirus (white bar) or a DN Ikaros expressing retrovirus (black bar). To standardize for slightly different starting frequencies, values are standardized relative to day 1 input values for either control or DN Ikaros transduction efficiencies with data showing mean ± SEM with 3–10 mice for control transduced and 6–10 mice for DN Ikaros transduced cells. Mice were bled on day 1, 7, and 39 post-transfer. Statistically significant differences (p<0.05) are indicated, calculated by unpaired t test. (B) Fold change in the abundance of retrovirally transduced cells in C57BL/6J recipient mice from panel A (either control or DN Ikaros transduced) in the blood (defined as the percentage of live, CD8+ MHC class II negative cells that were CD45.1+ Thy1.1+) following injection of either vehicle alone or recombinant IL-12 (1 µg) at day 162 and 165 post-adoptive transfer, with abundance post-treatment measured at day 168 post-transfer. For these studies, at day 1 after adoptive transfer, the frequency of adoptively transferred, retrovirally transduced cells (defined as CD45.1+ Thy1.1+) among CD8 T cells in the peripheral blood was 1.21±0.07% for control and 1.70±0.09% for DN Ikaros transduced cells (mean ± SEM). Among adoptively transferred cells (CD45.1+), the percentage (mean ± SEM) of transduced cells was 42.1±0.7% for control and 49.6±0.5% for DN Ikaros.

Mentions: To begin to address the in vivo consequence of DN Ikaros expression on CD8 T cell responses, we adoptively transferred DN Ikaros-transduced cells, grown in vitro, into normal C57BL/6J mice and followed their relative competitive ability in vivo (Fig. 6A). By monitoring the relative abundance of transduced cells in peripheral blood, standardized to input frequencies, we found that DN Ikaros-transduced cells had increased abundance relative to control-transduced cells at both day 7 and day 39 post-transfer (Fig. 6A). This initial advantage was not retained long-term, however, with equivalent abundance of DN Ikaros and control-transduced cells by day 160 post-transfer (not shown).


The Ikaros transcription factor regulates responsiveness to IL-12 and expression of IL-2 receptor alpha in mature, activated CD8 T cells.

Clambey ET, Collins B, Young MH, Eberlein J, David A, Kappler JW, Marrack P - PLoS ONE (2013)

CD8 T cells expressing dominant negative Ikaros have a modest advantage in vivo.(A) Relative abundance of retrovirally transduced cells within a pool of adoptively transferred CD8 T cells containing both transduced and non-transduced cells. Adoptively transferred cells were identified as live, CD8+ MHC class II negative cells that expressed CD45.1, with retrovirally transduced cells further expressing Thy1.1, with abundance measured within the peripheral blood of C57BL/6 recipient mice (which do not express CD45.1 or Thy1.1). Cells were transduced with a control retrovirus (white bar) or a DN Ikaros expressing retrovirus (black bar). To standardize for slightly different starting frequencies, values are standardized relative to day 1 input values for either control or DN Ikaros transduction efficiencies with data showing mean ± SEM with 3–10 mice for control transduced and 6–10 mice for DN Ikaros transduced cells. Mice were bled on day 1, 7, and 39 post-transfer. Statistically significant differences (p<0.05) are indicated, calculated by unpaired t test. (B) Fold change in the abundance of retrovirally transduced cells in C57BL/6J recipient mice from panel A (either control or DN Ikaros transduced) in the blood (defined as the percentage of live, CD8+ MHC class II negative cells that were CD45.1+ Thy1.1+) following injection of either vehicle alone or recombinant IL-12 (1 µg) at day 162 and 165 post-adoptive transfer, with abundance post-treatment measured at day 168 post-transfer. For these studies, at day 1 after adoptive transfer, the frequency of adoptively transferred, retrovirally transduced cells (defined as CD45.1+ Thy1.1+) among CD8 T cells in the peripheral blood was 1.21±0.07% for control and 1.70±0.09% for DN Ikaros transduced cells (mean ± SEM). Among adoptively transferred cells (CD45.1+), the percentage (mean ± SEM) of transduced cells was 42.1±0.7% for control and 49.6±0.5% for DN Ikaros.
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Related In: Results  -  Collection

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pone-0057435-g006: CD8 T cells expressing dominant negative Ikaros have a modest advantage in vivo.(A) Relative abundance of retrovirally transduced cells within a pool of adoptively transferred CD8 T cells containing both transduced and non-transduced cells. Adoptively transferred cells were identified as live, CD8+ MHC class II negative cells that expressed CD45.1, with retrovirally transduced cells further expressing Thy1.1, with abundance measured within the peripheral blood of C57BL/6 recipient mice (which do not express CD45.1 or Thy1.1). Cells were transduced with a control retrovirus (white bar) or a DN Ikaros expressing retrovirus (black bar). To standardize for slightly different starting frequencies, values are standardized relative to day 1 input values for either control or DN Ikaros transduction efficiencies with data showing mean ± SEM with 3–10 mice for control transduced and 6–10 mice for DN Ikaros transduced cells. Mice were bled on day 1, 7, and 39 post-transfer. Statistically significant differences (p<0.05) are indicated, calculated by unpaired t test. (B) Fold change in the abundance of retrovirally transduced cells in C57BL/6J recipient mice from panel A (either control or DN Ikaros transduced) in the blood (defined as the percentage of live, CD8+ MHC class II negative cells that were CD45.1+ Thy1.1+) following injection of either vehicle alone or recombinant IL-12 (1 µg) at day 162 and 165 post-adoptive transfer, with abundance post-treatment measured at day 168 post-transfer. For these studies, at day 1 after adoptive transfer, the frequency of adoptively transferred, retrovirally transduced cells (defined as CD45.1+ Thy1.1+) among CD8 T cells in the peripheral blood was 1.21±0.07% for control and 1.70±0.09% for DN Ikaros transduced cells (mean ± SEM). Among adoptively transferred cells (CD45.1+), the percentage (mean ± SEM) of transduced cells was 42.1±0.7% for control and 49.6±0.5% for DN Ikaros.
Mentions: To begin to address the in vivo consequence of DN Ikaros expression on CD8 T cell responses, we adoptively transferred DN Ikaros-transduced cells, grown in vitro, into normal C57BL/6J mice and followed their relative competitive ability in vivo (Fig. 6A). By monitoring the relative abundance of transduced cells in peripheral blood, standardized to input frequencies, we found that DN Ikaros-transduced cells had increased abundance relative to control-transduced cells at both day 7 and day 39 post-transfer (Fig. 6A). This initial advantage was not retained long-term, however, with equivalent abundance of DN Ikaros and control-transduced cells by day 160 post-transfer (not shown).

Bottom Line: While DN Ikaros-transduced cells had higher expression of the IL-2 receptor alpha chain, DN Ikaros-transduced cells achieved their competitive advantage through an IL-2 independent mechanism.Finally, the competitive advantage of DN Ikaros-transduced cells was manifested in vivo, following adoptive transfer of transduced cells.These data identify the Ikaros family of transcription factors as regulators of cytokine responsiveness in activated CD8 T cells, and suggest a role for this family in influencing effector and memory CD8 T cell differentiation.

View Article: PubMed Central - PubMed

Affiliation: Integrated Department of Immunology, University of Colorado Anschutz Medical Campus, Aurora, Colorado, USA. eric.clambey@ucdenver.edu

ABSTRACT
The Ikaros family of transcription factors is critical for normal T cell development while limiting malignant transformation. Mature CD8 T cells express multiple Ikaros family members, yet little is known about their function in this context. To test the functions of this gene family, we used retroviral transduction to express a naturally occurring, dominant negative (DN) isoform of Ikaros in activated CD8 T cells. Notably, expression of DN Ikaros profoundly enhanced the competitive advantage of activated CD8 T cells cultured in IL-12, such that by 6 days of culture, DN Ikaros-transduced cells were 100-fold more abundant than control cells. Expression of a DN isoform of Helios, a related Ikaros-family transcription factor, conferred a similar advantage to transduced cells in IL-12. While DN Ikaros-transduced cells had higher expression of the IL-2 receptor alpha chain, DN Ikaros-transduced cells achieved their competitive advantage through an IL-2 independent mechanism. Finally, the competitive advantage of DN Ikaros-transduced cells was manifested in vivo, following adoptive transfer of transduced cells. These data identify the Ikaros family of transcription factors as regulators of cytokine responsiveness in activated CD8 T cells, and suggest a role for this family in influencing effector and memory CD8 T cell differentiation.

Show MeSH
Related in: MedlinePlus