Expression profile of peripheral tissue antigen genes in medullary thymic epithelial cells (mTECs) is dependent on mRNA levels of autoimmune regulator (Aire)

Oliveira, Érika Arantes de; Macedo, Cristiana Santos de; Donate, Paula B.; Almeida, Renata Servan de; Pezzi, Nicole; Nguyen, Catherine; Rossi, Marcos A.; Sakamoto-Hojo, Elza T.; Donadi, Eduardo Antônio; Passos, Geraldo A. S.

doi:10.1016/j.imbio.2012.02.005

Cited by 23 publications

(25 citation statements)

References 45 publications

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“…A 0.33-mm-diameter injection needle was connected to the crocodile clip corresponding to the anode and then introduced between the first and second ribs where the thymus was located at a 45° angle from the longitudinal axis. As previously reported (15), we observed that inserting the needle to a 5-mm depth was necessary and sufficient to reach the thymus without affecting proximal vital organs such as the heart and lungs.…”

Section: Methodssupporting

confidence: 81%

“…We used the TriFecta (IDT, Integrated DNA Technologies, Coralville, IA, USA) anti-Aire siRNA sequence (GGAUUCUCUUUAAG-GACUACAAUCTAGAUUGUAGUCCUUAAAGAGAAUCCUC) to knock down the Aire mRNA in vivo by thymic electro-transfection of BALB/c mice as previously described (15, 39). Briefly, the mice were anesthetized by intraperitoneal injection with a mixture of ketamine (Sigma-Aldrich K-003) (100 mg/kg body weight) and xylazine (Sigma-Aldrich X1251) (10 mg/kg body weight).…”

Section: Methodsmentioning

confidence: 99%

“…The results showed that the thymus tissue structure was preserved within the evaluated time frame (48 h post-electro-transfection) (15). …”

Section: Methodsmentioning

confidence: 99%

“…The thymuses were dissected 48 h post-electro-transfection, and the thymic stroma was separated as previously described (15, 40). Briefly, the thymuses were dissected and trimmed of fat and connective tissue.…”

Section: Methodsmentioning

confidence: 99%

“…Previously, we demonstrated that slight variations in wild-type thymic Aire gene expression in vivo were sufficient to affect the expression of AIRE-dependent PTAs, either spontaneously as observed in the non-obese diabetic (NOD) mouse strain (14) or following induction by gene knockdown either in vivo in the thymuses of BALB/c strain mice or in vitro in a mTEC cell line (15). …”

Section: Introductionmentioning

confidence: 99%

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

et al. 2016

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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.

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Section: Methodssupporting

confidence: 81%

Section: Methodsmentioning

confidence: 99%

“…The results showed that the thymus tissue structure was preserved within the evaluated time frame (48 h post-electro-transfection) (15). …”

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%