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Primary murine CD4+ T cells fail to acquire the ability to produce effector cytokines when active Ras is present during Th1/Th2 differentiation.

Janardhan SV, Marks R, Gajewski TF - PLoS ONE (2014)

Bottom Line: This differentiation defect was not due to deficient STAT4 or STAT6 activation by IL-12 or IL-4, respectively, nor was it associated with deficient induction of T-bet and GATA-3 expression.Impaired effector cytokine production in active Ras-transduced cells was associated with deficient demethylation of the IL-4 gene locus.Our results indicate that, despite augmenting acute activation of naïve T cells, constitutive Ras signaling inhibits the ability of CD4+ T cells to properly differentiate into Th1/Th2 effector cytokine-producing cells, in part by interfering with epigenetic modification of effector gene loci.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, The University of Chicago, Chicago, Illinois, United States of America.

ABSTRACT
Constitutive Ras signaling has been shown to augment IL-2 production, reverse anergy, and functionally replace many aspects of CD28 co-stimulation in CD4+ T cells. These data raise the possibility that introduction of active Ras into primary T cells might result in improved functionality in pathologic situations of T cell dysfunction, such as cancer or chronic viral infection. To test the biologic effects of active Ras in primary T cells, CD4+ T cells from Coxsackie-Adenovirus Receptor Transgenic mice were transduced with an adenovirus encoding active Ras. As expected, active Ras augmented IL-2 production in naive CD4+ T cells. However, when cells were cultured for 4 days under conditions to promote effector cell differentiation, active Ras inhibited the ability of CD4+ T cells to acquire a Th1 or Th2 effector cytokine profile. This differentiation defect was not due to deficient STAT4 or STAT6 activation by IL-12 or IL-4, respectively, nor was it associated with deficient induction of T-bet and GATA-3 expression. Impaired effector cytokine production in active Ras-transduced cells was associated with deficient demethylation of the IL-4 gene locus. Our results indicate that, despite augmenting acute activation of naïve T cells, constitutive Ras signaling inhibits the ability of CD4+ T cells to properly differentiate into Th1/Th2 effector cytokine-producing cells, in part by interfering with epigenetic modification of effector gene loci. Alternative strategies to potentiate Ras pathway signaling in T cells in a more regulated fashion should be considered as a therapeutic approach to improve immune responses in vivo.

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Model of CD4+ T cell activation and differentiation in the presence of constitutive Ras signaling.Initial activation of naive T cells induces Ras activation that persists for several hours until productive signaling culminates in the targets of acute activation (e.g. IL-2 transcription). Constitutive Ras signaling in Ras61L-transduced cells augments signaling events during this time, resulting in augmented acute activation and IL-2 production. After these acute activation events have been reached in normal cells, regulatory mechanisms cause Ras signaling pathways to become attenuated/regulated. This allows for robust differentiation including the ability to signal through polarizing cytokine receptors, induce lineage-specific transcription factors, proliferate and remodel effector cytokine loci. Cells that maintain deregulated Ras signaling are able to acquire several aspects of differentiation listed above, but are unable to produce effector cytokines. This occurs even in cells that are able to overcome a partial inhibition of proliferation, and is associated with persistent methylation of the effector cytokine locus.
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pone-0112831-g007: Model of CD4+ T cell activation and differentiation in the presence of constitutive Ras signaling.Initial activation of naive T cells induces Ras activation that persists for several hours until productive signaling culminates in the targets of acute activation (e.g. IL-2 transcription). Constitutive Ras signaling in Ras61L-transduced cells augments signaling events during this time, resulting in augmented acute activation and IL-2 production. After these acute activation events have been reached in normal cells, regulatory mechanisms cause Ras signaling pathways to become attenuated/regulated. This allows for robust differentiation including the ability to signal through polarizing cytokine receptors, induce lineage-specific transcription factors, proliferate and remodel effector cytokine loci. Cells that maintain deregulated Ras signaling are able to acquire several aspects of differentiation listed above, but are unable to produce effector cytokines. This occurs even in cells that are able to overcome a partial inhibition of proliferation, and is associated with persistent methylation of the effector cytokine locus.

Mentions: It is of interest that deregulated Ras signaling did not lead to a complete failure of the process of differentiation, as several other aspects of Th1/Th2 differentiation, including responsiveness to IL-12 and IL-4 cytokine signaling and upregulated expression of T-bet and GATA-3, proceeded normally. While active Ras did induce a partial anti-proliferative effect, this could not fully explain the effect of Ras on impaired cytokine production. The inability of Ras61L-transduced cells to properly remodel the IL-4 locus suggests that epigenetic modification during T cell differentiation requires properly regulated Ras signaling. Ras61L-transduced cells not only failed to upregulate effector cytokines, but also failed to modulate IL-2 production in a lineage-specific way, thus maintaining a more “naïve-like” cytokine profile. This observation suggests a global effect on cytokine regulation that may be consistent with a failure of epigenetic modifications. A model of this process is shown pictorially in Figure 7.


Primary murine CD4+ T cells fail to acquire the ability to produce effector cytokines when active Ras is present during Th1/Th2 differentiation.

Janardhan SV, Marks R, Gajewski TF - PLoS ONE (2014)

Model of CD4+ T cell activation and differentiation in the presence of constitutive Ras signaling.Initial activation of naive T cells induces Ras activation that persists for several hours until productive signaling culminates in the targets of acute activation (e.g. IL-2 transcription). Constitutive Ras signaling in Ras61L-transduced cells augments signaling events during this time, resulting in augmented acute activation and IL-2 production. After these acute activation events have been reached in normal cells, regulatory mechanisms cause Ras signaling pathways to become attenuated/regulated. This allows for robust differentiation including the ability to signal through polarizing cytokine receptors, induce lineage-specific transcription factors, proliferate and remodel effector cytokine loci. Cells that maintain deregulated Ras signaling are able to acquire several aspects of differentiation listed above, but are unable to produce effector cytokines. This occurs even in cells that are able to overcome a partial inhibition of proliferation, and is associated with persistent methylation of the effector cytokine locus.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4232516&req=5

pone-0112831-g007: Model of CD4+ T cell activation and differentiation in the presence of constitutive Ras signaling.Initial activation of naive T cells induces Ras activation that persists for several hours until productive signaling culminates in the targets of acute activation (e.g. IL-2 transcription). Constitutive Ras signaling in Ras61L-transduced cells augments signaling events during this time, resulting in augmented acute activation and IL-2 production. After these acute activation events have been reached in normal cells, regulatory mechanisms cause Ras signaling pathways to become attenuated/regulated. This allows for robust differentiation including the ability to signal through polarizing cytokine receptors, induce lineage-specific transcription factors, proliferate and remodel effector cytokine loci. Cells that maintain deregulated Ras signaling are able to acquire several aspects of differentiation listed above, but are unable to produce effector cytokines. This occurs even in cells that are able to overcome a partial inhibition of proliferation, and is associated with persistent methylation of the effector cytokine locus.
Mentions: It is of interest that deregulated Ras signaling did not lead to a complete failure of the process of differentiation, as several other aspects of Th1/Th2 differentiation, including responsiveness to IL-12 and IL-4 cytokine signaling and upregulated expression of T-bet and GATA-3, proceeded normally. While active Ras did induce a partial anti-proliferative effect, this could not fully explain the effect of Ras on impaired cytokine production. The inability of Ras61L-transduced cells to properly remodel the IL-4 locus suggests that epigenetic modification during T cell differentiation requires properly regulated Ras signaling. Ras61L-transduced cells not only failed to upregulate effector cytokines, but also failed to modulate IL-2 production in a lineage-specific way, thus maintaining a more “naïve-like” cytokine profile. This observation suggests a global effect on cytokine regulation that may be consistent with a failure of epigenetic modifications. A model of this process is shown pictorially in Figure 7.

Bottom Line: This differentiation defect was not due to deficient STAT4 or STAT6 activation by IL-12 or IL-4, respectively, nor was it associated with deficient induction of T-bet and GATA-3 expression.Impaired effector cytokine production in active Ras-transduced cells was associated with deficient demethylation of the IL-4 gene locus.Our results indicate that, despite augmenting acute activation of naïve T cells, constitutive Ras signaling inhibits the ability of CD4+ T cells to properly differentiate into Th1/Th2 effector cytokine-producing cells, in part by interfering with epigenetic modification of effector gene loci.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, The University of Chicago, Chicago, Illinois, United States of America.

ABSTRACT
Constitutive Ras signaling has been shown to augment IL-2 production, reverse anergy, and functionally replace many aspects of CD28 co-stimulation in CD4+ T cells. These data raise the possibility that introduction of active Ras into primary T cells might result in improved functionality in pathologic situations of T cell dysfunction, such as cancer or chronic viral infection. To test the biologic effects of active Ras in primary T cells, CD4+ T cells from Coxsackie-Adenovirus Receptor Transgenic mice were transduced with an adenovirus encoding active Ras. As expected, active Ras augmented IL-2 production in naive CD4+ T cells. However, when cells were cultured for 4 days under conditions to promote effector cell differentiation, active Ras inhibited the ability of CD4+ T cells to acquire a Th1 or Th2 effector cytokine profile. This differentiation defect was not due to deficient STAT4 or STAT6 activation by IL-12 or IL-4, respectively, nor was it associated with deficient induction of T-bet and GATA-3 expression. Impaired effector cytokine production in active Ras-transduced cells was associated with deficient demethylation of the IL-4 gene locus. Our results indicate that, despite augmenting acute activation of naïve T cells, constitutive Ras signaling inhibits the ability of CD4+ T cells to properly differentiate into Th1/Th2 effector cytokine-producing cells, in part by interfering with epigenetic modification of effector gene loci. Alternative strategies to potentiate Ras pathway signaling in T cells in a more regulated fashion should be considered as a therapeutic approach to improve immune responses in vivo.

Show MeSH
Related in: MedlinePlus