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miR-20a inhibits TCR-mediated signaling and cytokine production in human naïve CD4+ T cells.

Reddycherla AV, Meinert I, Reinhold A, Reinhold D, Schraven B, Simeoni L - PLoS ONE (2015)

Bottom Line: In this study, we have focused our investigation on miR-20a, a member of the miR-17-92 cluster, whose expression is decreased in patients suffering from multiple sclerosis.We have found that miR-20a is rapidly induced upon TCR-triggering in primary human naïve CD4+ T cells and that its transcription is regulated in a Erk-, NF-κB-, and Ca++-dependent manner.However, miR-20a overexpression strongly suppresses IL-10 secretion and moderately decreases IL-2, IL-6 and IL8 production, which are crucial regulators of inflammatory responses.

View Article: PubMed Central - PubMed

Affiliation: Otto-von-Guericke University, Institute of Molecular and Clinical Immunology, Leipziger Str. 44, Magdeburg, Germany.

ABSTRACT
Upon TCR stimulation by peptide-MHC complexes, CD4+ T cells undergo activation and proliferation. This process will ultimately culminate in T-cell differentiation and the acquisition of effector functions. The production of specific cytokines by differentiated CD4+ T cells is crucial for the generation of the appropriate immune response. Altered CD4+ T-cell activation and cytokine production result in chronic inflammatory conditions and autoimmune disorders. miRNAs have been shown to be important regulators of T-cell biology. In this study, we have focused our investigation on miR-20a, a member of the miR-17-92 cluster, whose expression is decreased in patients suffering from multiple sclerosis. We have found that miR-20a is rapidly induced upon TCR-triggering in primary human naïve CD4+ T cells and that its transcription is regulated in a Erk-, NF-κB-, and Ca++-dependent manner. We have further shown that overexpression of miR-20a inhibits TCR-mediated signaling but not the proliferation of primary human naïve CD4+ T cells. However, miR-20a overexpression strongly suppresses IL-10 secretion and moderately decreases IL-2, IL-6 and IL8 production, which are crucial regulators of inflammatory responses. Our study suggests that miR-20a is a new player in the regulation of TCR signaling strength and cytokine production.

No MeSH data available.


Related in: MedlinePlus

miR-20a does not affect the expression of signaling molecules and CD3 and CD28 receptors.(A) Lysates from T cells transfected with either miR-20a or miR-control were prepared and analyzed by immunoblotting using the indicated Abs. (B) CD4+ T cells overexpressing either miR-20a or miR-control cells were stained with antibodies against CD3 and CD28 and analyzed by flow cytometry. Histograms overlay show the expression of CD3 (left) and CD28 (right) in miR-20a versus miR-control transfected cells.
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pone.0125311.g003: miR-20a does not affect the expression of signaling molecules and CD3 and CD28 receptors.(A) Lysates from T cells transfected with either miR-20a or miR-control were prepared and analyzed by immunoblotting using the indicated Abs. (B) CD4+ T cells overexpressing either miR-20a or miR-control cells were stained with antibodies against CD3 and CD28 and analyzed by flow cytometry. Histograms overlay show the expression of CD3 (left) and CD28 (right) in miR-20a versus miR-control transfected cells.

Mentions: To exclude the possibility that the overexpression of miR-20a affects the expression of crucial signaling molecules, we performed additional experiments. We have found that overexpression of miR-20a does not affect the expression of Lck, Zap-70, LAT, PLC-γ, SLP-76, GADS, Sos1, Erk in resting (Fig 3A) as well as in activated CD4+ T cells (data not shown). Furthermore, there is also no difference observed in the expression levels of BIM, PTEN and STAT3 that have been previously shown to be the targets of the miR-17-92 cluster in both resting (Fig 3A) and activated CD4+ T cells (data not shown). Moreover, to exclude the possibility that defective TCR signaling is due to reduced receptor levels, we also examined the expression of CD3 and CD28. FACS analyses revealed that miR-20a overexpression does not change the densities of CD3 and CD28 on the cell surface compared to control (Fig 3B).


miR-20a inhibits TCR-mediated signaling and cytokine production in human naïve CD4+ T cells.

Reddycherla AV, Meinert I, Reinhold A, Reinhold D, Schraven B, Simeoni L - PLoS ONE (2015)

miR-20a does not affect the expression of signaling molecules and CD3 and CD28 receptors.(A) Lysates from T cells transfected with either miR-20a or miR-control were prepared and analyzed by immunoblotting using the indicated Abs. (B) CD4+ T cells overexpressing either miR-20a or miR-control cells were stained with antibodies against CD3 and CD28 and analyzed by flow cytometry. Histograms overlay show the expression of CD3 (left) and CD28 (right) in miR-20a versus miR-control transfected cells.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4401545&req=5

pone.0125311.g003: miR-20a does not affect the expression of signaling molecules and CD3 and CD28 receptors.(A) Lysates from T cells transfected with either miR-20a or miR-control were prepared and analyzed by immunoblotting using the indicated Abs. (B) CD4+ T cells overexpressing either miR-20a or miR-control cells were stained with antibodies against CD3 and CD28 and analyzed by flow cytometry. Histograms overlay show the expression of CD3 (left) and CD28 (right) in miR-20a versus miR-control transfected cells.
Mentions: To exclude the possibility that the overexpression of miR-20a affects the expression of crucial signaling molecules, we performed additional experiments. We have found that overexpression of miR-20a does not affect the expression of Lck, Zap-70, LAT, PLC-γ, SLP-76, GADS, Sos1, Erk in resting (Fig 3A) as well as in activated CD4+ T cells (data not shown). Furthermore, there is also no difference observed in the expression levels of BIM, PTEN and STAT3 that have been previously shown to be the targets of the miR-17-92 cluster in both resting (Fig 3A) and activated CD4+ T cells (data not shown). Moreover, to exclude the possibility that defective TCR signaling is due to reduced receptor levels, we also examined the expression of CD3 and CD28. FACS analyses revealed that miR-20a overexpression does not change the densities of CD3 and CD28 on the cell surface compared to control (Fig 3B).

Bottom Line: In this study, we have focused our investigation on miR-20a, a member of the miR-17-92 cluster, whose expression is decreased in patients suffering from multiple sclerosis.We have found that miR-20a is rapidly induced upon TCR-triggering in primary human naïve CD4+ T cells and that its transcription is regulated in a Erk-, NF-κB-, and Ca++-dependent manner.However, miR-20a overexpression strongly suppresses IL-10 secretion and moderately decreases IL-2, IL-6 and IL8 production, which are crucial regulators of inflammatory responses.

View Article: PubMed Central - PubMed

Affiliation: Otto-von-Guericke University, Institute of Molecular and Clinical Immunology, Leipziger Str. 44, Magdeburg, Germany.

ABSTRACT
Upon TCR stimulation by peptide-MHC complexes, CD4+ T cells undergo activation and proliferation. This process will ultimately culminate in T-cell differentiation and the acquisition of effector functions. The production of specific cytokines by differentiated CD4+ T cells is crucial for the generation of the appropriate immune response. Altered CD4+ T-cell activation and cytokine production result in chronic inflammatory conditions and autoimmune disorders. miRNAs have been shown to be important regulators of T-cell biology. In this study, we have focused our investigation on miR-20a, a member of the miR-17-92 cluster, whose expression is decreased in patients suffering from multiple sclerosis. We have found that miR-20a is rapidly induced upon TCR-triggering in primary human naïve CD4+ T cells and that its transcription is regulated in a Erk-, NF-κB-, and Ca++-dependent manner. We have further shown that overexpression of miR-20a inhibits TCR-mediated signaling but not the proliferation of primary human naïve CD4+ T cells. However, miR-20a overexpression strongly suppresses IL-10 secretion and moderately decreases IL-2, IL-6 and IL8 production, which are crucial regulators of inflammatory responses. Our study suggests that miR-20a is a new player in the regulation of TCR signaling strength and cytokine production.

No MeSH data available.


Related in: MedlinePlus