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The miR-17 ∼ 92a cluster of microRNAs is required for the fitness of Foxp3+ regulatory T cells.

Skinner JP, Keown AA, Chong MM - PLoS ONE (2014)

Bottom Line: In this study, we identified members of the miR-17 ∼ 92a cluster of microRNAs to be enriched in regulatory T cells.This led to a loss of Foxp3(+) cells over time, particularly in competitive settings, and culminated in a range of immunologic perturbations.Thus, miR-17 ∼ 92a-target interactions are part of the essential microRNA networks that safeguard the regulatory T cell lineage.

View Article: PubMed Central - PubMed

Affiliation: St Vincent's Institute of Medical Research, Fitzroy, Victoria, Australia.

ABSTRACT
By genetic inactivation of key microRNA biogenesis enzymes, we and others have previously demonstrated the critical requirement of the microRNA pathway for the differentiation and function of Foxp3(+) regulatory T cells. In this study, we identified members of the miR-17 ∼ 92a cluster of microRNAs to be enriched in regulatory T cells. To investigate the function of this microRNA cluster, we deleted the gene specifically in Foxp3(+) cells in mice. We found that miR-17 ∼ 92a is required for the fitness of regulatory T cells, and deficiency impacted at the level of apoptosis and proliferation of these cells. This led to a loss of Foxp3(+) cells over time, particularly in competitive settings, and culminated in a range of immunologic perturbations. Thus, miR-17 ∼ 92a-target interactions are part of the essential microRNA networks that safeguard the regulatory T cell lineage.

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Older mice with Treg-specific miR-17∼92a deficiency display perturbations in Treg populations.(A) Foxp3+ (YFP+) cells were sorted from the spleens of Mir17∼92afl/fl Foxp3CreYFP/CreYFP(Y) (KO) and Mir17∼92afl/+ Foxp3CreYFP/CreYFP(Y) control (WT) mice, then analyzed for miR-17 expression by Taqman-based RT-PCR. As a negative control, miR-21 expression was also measured. All miRNA levels were normalized to U6 snRNA. Frequency of Foxp3+ Tregs in the (B) spleen and (C) lamina propria of the colon. The data is expressed as a percentage of total CD4+ T cells. Littermate KO and WT pairs at ∼5 weeks (n = 5), ∼15 weeks (n = 6–8) and ∼50 weeks (n = 9) of age were analyzed. Paired t tests were performed for statistical comparisons. Only significant differences are indicated. (D) Examples of Treg profiles of mouse pairs at 15 and 50 weeks of age. Shown is the colonic lamina propria gated on TCRβ+ cells. Note that all TCRβ+ cells are CD4+.
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pone-0088997-g002: Older mice with Treg-specific miR-17∼92a deficiency display perturbations in Treg populations.(A) Foxp3+ (YFP+) cells were sorted from the spleens of Mir17∼92afl/fl Foxp3CreYFP/CreYFP(Y) (KO) and Mir17∼92afl/+ Foxp3CreYFP/CreYFP(Y) control (WT) mice, then analyzed for miR-17 expression by Taqman-based RT-PCR. As a negative control, miR-21 expression was also measured. All miRNA levels were normalized to U6 snRNA. Frequency of Foxp3+ Tregs in the (B) spleen and (C) lamina propria of the colon. The data is expressed as a percentage of total CD4+ T cells. Littermate KO and WT pairs at ∼5 weeks (n = 5), ∼15 weeks (n = 6–8) and ∼50 weeks (n = 9) of age were analyzed. Paired t tests were performed for statistical comparisons. Only significant differences are indicated. (D) Examples of Treg profiles of mouse pairs at 15 and 50 weeks of age. Shown is the colonic lamina propria gated on TCRβ+ cells. Note that all TCRβ+ cells are CD4+.

Mentions: The miR-17∼92 cluster of miRNAs are all derived from a single polycistronic transcript driven by a single promoter [32], [33]. To investigate the requirement of this miRNA cluster in Tregs, we generated Treg-specific miR-17∼92a deficient mice by intercrossing mice with a LoxP-flanked Mir17∼92afl allele with a CreYFP allele expressed from the Foxp3 locus. This resulted in a loss of miR-17∼92a cluster miRNAs but not other miRNAs (Figure 2A). In agreement with a recent report [34], Treg numbers appeared normal in the secondary lymphoid organs of young KO mice (Figure 2B). However, perturbations in Treg populations became apparent as the mice aged. Treg frequency was significantly reduced in the spleens of 50 week of KO mice. Even more dramatic was the reduction in the lamina propria of the colon. Large numbers of Tregs are normally found in the lamina propria of the colon as a result of interactions with commensal flora [9]. However, a significant reduction in Treg numbers was already evident in 15 week KO mice (Figure 2C and D).


The miR-17 ∼ 92a cluster of microRNAs is required for the fitness of Foxp3+ regulatory T cells.

Skinner JP, Keown AA, Chong MM - PLoS ONE (2014)

Older mice with Treg-specific miR-17∼92a deficiency display perturbations in Treg populations.(A) Foxp3+ (YFP+) cells were sorted from the spleens of Mir17∼92afl/fl Foxp3CreYFP/CreYFP(Y) (KO) and Mir17∼92afl/+ Foxp3CreYFP/CreYFP(Y) control (WT) mice, then analyzed for miR-17 expression by Taqman-based RT-PCR. As a negative control, miR-21 expression was also measured. All miRNA levels were normalized to U6 snRNA. Frequency of Foxp3+ Tregs in the (B) spleen and (C) lamina propria of the colon. The data is expressed as a percentage of total CD4+ T cells. Littermate KO and WT pairs at ∼5 weeks (n = 5), ∼15 weeks (n = 6–8) and ∼50 weeks (n = 9) of age were analyzed. Paired t tests were performed for statistical comparisons. Only significant differences are indicated. (D) Examples of Treg profiles of mouse pairs at 15 and 50 weeks of age. Shown is the colonic lamina propria gated on TCRβ+ cells. Note that all TCRβ+ cells are CD4+.
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pone-0088997-g002: Older mice with Treg-specific miR-17∼92a deficiency display perturbations in Treg populations.(A) Foxp3+ (YFP+) cells were sorted from the spleens of Mir17∼92afl/fl Foxp3CreYFP/CreYFP(Y) (KO) and Mir17∼92afl/+ Foxp3CreYFP/CreYFP(Y) control (WT) mice, then analyzed for miR-17 expression by Taqman-based RT-PCR. As a negative control, miR-21 expression was also measured. All miRNA levels were normalized to U6 snRNA. Frequency of Foxp3+ Tregs in the (B) spleen and (C) lamina propria of the colon. The data is expressed as a percentage of total CD4+ T cells. Littermate KO and WT pairs at ∼5 weeks (n = 5), ∼15 weeks (n = 6–8) and ∼50 weeks (n = 9) of age were analyzed. Paired t tests were performed for statistical comparisons. Only significant differences are indicated. (D) Examples of Treg profiles of mouse pairs at 15 and 50 weeks of age. Shown is the colonic lamina propria gated on TCRβ+ cells. Note that all TCRβ+ cells are CD4+.
Mentions: The miR-17∼92 cluster of miRNAs are all derived from a single polycistronic transcript driven by a single promoter [32], [33]. To investigate the requirement of this miRNA cluster in Tregs, we generated Treg-specific miR-17∼92a deficient mice by intercrossing mice with a LoxP-flanked Mir17∼92afl allele with a CreYFP allele expressed from the Foxp3 locus. This resulted in a loss of miR-17∼92a cluster miRNAs but not other miRNAs (Figure 2A). In agreement with a recent report [34], Treg numbers appeared normal in the secondary lymphoid organs of young KO mice (Figure 2B). However, perturbations in Treg populations became apparent as the mice aged. Treg frequency was significantly reduced in the spleens of 50 week of KO mice. Even more dramatic was the reduction in the lamina propria of the colon. Large numbers of Tregs are normally found in the lamina propria of the colon as a result of interactions with commensal flora [9]. However, a significant reduction in Treg numbers was already evident in 15 week KO mice (Figure 2C and D).

Bottom Line: In this study, we identified members of the miR-17 ∼ 92a cluster of microRNAs to be enriched in regulatory T cells.This led to a loss of Foxp3(+) cells over time, particularly in competitive settings, and culminated in a range of immunologic perturbations.Thus, miR-17 ∼ 92a-target interactions are part of the essential microRNA networks that safeguard the regulatory T cell lineage.

View Article: PubMed Central - PubMed

Affiliation: St Vincent's Institute of Medical Research, Fitzroy, Victoria, Australia.

ABSTRACT
By genetic inactivation of key microRNA biogenesis enzymes, we and others have previously demonstrated the critical requirement of the microRNA pathway for the differentiation and function of Foxp3(+) regulatory T cells. In this study, we identified members of the miR-17 ∼ 92a cluster of microRNAs to be enriched in regulatory T cells. To investigate the function of this microRNA cluster, we deleted the gene specifically in Foxp3(+) cells in mice. We found that miR-17 ∼ 92a is required for the fitness of regulatory T cells, and deficiency impacted at the level of apoptosis and proliferation of these cells. This led to a loss of Foxp3(+) cells over time, particularly in competitive settings, and culminated in a range of immunologic perturbations. Thus, miR-17 ∼ 92a-target interactions are part of the essential microRNA networks that safeguard the regulatory T cell lineage.

Show MeSH
Related in: MedlinePlus