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Aire Downregulation Is Associated with Changes in the Posttranscriptional Control of Peripheral Tissue Antigens in Medullary Thymic Epithelial Cells

View Article: PubMed Central - PubMed

ABSTRACT

Autoimmune regulator (Aire) is a transcriptional regulator of peripheral tissue antigens (PTAs) and microRNAs (miRNAs) in medullary thymic epithelial cells (mTECs). In this study, we tested the hypothesis that Aire also played a role as an upstream posttranscriptional controller in these cells and that variation in its expression might be associated with changes in the interactions between miRNAs and the mRNAs encoding PTAs. We demonstrated that downregulation of Aire in vivo in the thymuses of BALB/c mice imbalanced the large-scale expression of these two RNA species and consequently their interactions. The expression profiles of a large set of mTEC miRNAs and mRNAs isolated from the thymuses of mice subjected (or not) to small-interfering-induced Aire gene knockdown revealed that 87 miRNAs and 4,558 mRNAs were differentially expressed. The reconstruction of the miRNA–mRNA interaction networks demonstrated that interactions between these RNAs were under Aire influence and therefore changed when this gene was downregulated. Prior to Aire-knockdown, only members of the miR-let-7 family interacted with a set of PTA mRNAs. Under Aire-knockdown conditions, a larger set of miRNA families and their members established this type of interaction. Notably, no previously described Aire-dependent PTA interacted with the miRNAs, indicating that these PTAs were somehow refractory. The miRNA–mRNA interactions were validated by calculating the minimal free energy of the pairings between the miRNA seed regions and the mRNA 3′ UTRs and within the cellular milieu using the luciferase reporter gene assay. These results suggest the existence of a link between transcriptional and posttranscriptional control because Aire downregulation alters the miRNA–mRNA network controlling PTAs in mTEC cells.

No MeSH data available.


Posttranscriptional miRNA–mRNA interaction network based on microarray expression data. The network of control mTEC cells (A) shows the participation of six miRNAs of let-7 family that interact with PTA mRNAs targets. The highlighted interaction (let-7a–Hmga2 mRNA) was selected for further validations. The network of Aire-knockdown mTEC cells (B) shows the participation of 15 miRNAs from different families that interact with a restricted group of PTA mRNA targets. The highlighted interaction (miR-378–Prpsap1 mRNA) was selected for further validations. Interaction networks were reconstructed by using the GenMir++ algorithm, which consider the respective modulation of miRNAs (upregulated) and of mRNA targets (downregulated).
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Figure 5: Posttranscriptional miRNA–mRNA interaction network based on microarray expression data. The network of control mTEC cells (A) shows the participation of six miRNAs of let-7 family that interact with PTA mRNAs targets. The highlighted interaction (let-7a–Hmga2 mRNA) was selected for further validations. The network of Aire-knockdown mTEC cells (B) shows the participation of 15 miRNAs from different families that interact with a restricted group of PTA mRNA targets. The highlighted interaction (miR-378–Prpsap1 mRNA) was selected for further validations. Interaction networks were reconstructed by using the GenMir++ algorithm, which consider the respective modulation of miRNAs (upregulated) and of mRNA targets (downregulated).

Mentions: The microarray-normalized data were used as inputs for the GenMir++ algorithm to reconstruct two miRNA–mRNA interaction networks (Figures 5A,B). The thermodynamic hybridization stabilities of the selected miRNA–mRNA pairs were reanalyzed using the RNAhybrid algorithm, which calculated the MFE of annealing. We selected only miRNA–mRNA pairs with an MFE ≤−20 kcal/mol for the subsequent experiments.


Aire Downregulation Is Associated with Changes in the Posttranscriptional Control of Peripheral Tissue Antigens in Medullary Thymic Epithelial Cells
Posttranscriptional miRNA–mRNA interaction network based on microarray expression data. The network of control mTEC cells (A) shows the participation of six miRNAs of let-7 family that interact with PTA mRNAs targets. The highlighted interaction (let-7a–Hmga2 mRNA) was selected for further validations. The network of Aire-knockdown mTEC cells (B) shows the participation of 15 miRNAs from different families that interact with a restricted group of PTA mRNA targets. The highlighted interaction (miR-378–Prpsap1 mRNA) was selected for further validations. Interaction networks were reconstructed by using the GenMir++ algorithm, which consider the respective modulation of miRNAs (upregulated) and of mRNA targets (downregulated).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC5120147&req=5

Figure 5: Posttranscriptional miRNA–mRNA interaction network based on microarray expression data. The network of control mTEC cells (A) shows the participation of six miRNAs of let-7 family that interact with PTA mRNAs targets. The highlighted interaction (let-7a–Hmga2 mRNA) was selected for further validations. The network of Aire-knockdown mTEC cells (B) shows the participation of 15 miRNAs from different families that interact with a restricted group of PTA mRNA targets. The highlighted interaction (miR-378–Prpsap1 mRNA) was selected for further validations. Interaction networks were reconstructed by using the GenMir++ algorithm, which consider the respective modulation of miRNAs (upregulated) and of mRNA targets (downregulated).
Mentions: The microarray-normalized data were used as inputs for the GenMir++ algorithm to reconstruct two miRNA–mRNA interaction networks (Figures 5A,B). The thermodynamic hybridization stabilities of the selected miRNA–mRNA pairs were reanalyzed using the RNAhybrid algorithm, which calculated the MFE of annealing. We selected only miRNA–mRNA pairs with an MFE ≤−20 kcal/mol for the subsequent experiments.

View Article: PubMed Central - PubMed

ABSTRACT

Autoimmune regulator (Aire) is a transcriptional regulator of peripheral tissue antigens (PTAs) and microRNAs (miRNAs) in medullary thymic epithelial cells (mTECs). In this study, we tested the hypothesis that Aire also played a role as an upstream posttranscriptional controller in these cells and that variation in its expression might be associated with changes in the interactions between miRNAs and the mRNAs encoding PTAs. We demonstrated that downregulation of Aire in vivo in the thymuses of BALB/c mice imbalanced the large-scale expression of these two RNA species and consequently their interactions. The expression profiles of a large set of mTEC miRNAs and mRNAs isolated from the thymuses of mice subjected (or not) to small-interfering-induced Aire gene knockdown revealed that 87 miRNAs and 4,558 mRNAs were differentially expressed. The reconstruction of the miRNA–mRNA interaction networks demonstrated that interactions between these RNAs were under Aire influence and therefore changed when this gene was downregulated. Prior to Aire-knockdown, only members of the miR-let-7 family interacted with a set of PTA mRNAs. Under Aire-knockdown conditions, a larger set of miRNA families and their members established this type of interaction. Notably, no previously described Aire-dependent PTA interacted with the miRNAs, indicating that these PTAs were somehow refractory. The miRNA–mRNA interactions were validated by calculating the minimal free energy of the pairings between the miRNA seed regions and the mRNA 3′ UTRs and within the cellular milieu using the luciferase reporter gene assay. These results suggest the existence of a link between transcriptional and posttranscriptional control because Aire downregulation alters the miRNA–mRNA network controlling PTAs in mTEC cells.

No MeSH data available.