No abstract

miR-146a inhibits inflammatory responses in human keratinocytes and in different mouse models of skin inflammation. Little is known about the role of miR-146b in the skin. In the present study, we confirmed the increased expression of miR-146a and miR-146b (miR-146a/b) in lesional skin of psoriasis patients. The expression of miR-146a was about 2-fold higher than that of miR-146b in healthy human skin and it was more strongly induced by stimulation of pro-inflammatory cytokines in keratinocytes and fibroblasts. miR-146a/b target genes regulating inflammatory responses or proliferation were altered in the skin of psoriasis patients, among which FERMT1 was verified as direct target of miR-146a. In silico analysis of genome-wide data from >4,000 psoriasis cases and >8,000 controls confirmed a moderate association between psoriasis and genetic variants in miR-146a gene. Transfection of miR-146a/b suppressed and inhibition enhanced keratinocyte proliferation and the expression of psoriasis-related target genes. Enhanced expression of miR-146a/b-influenced genes was detected in cultured keratinocytes from miR-146a−/− and skin fibroblasts from miR-146a−/− and miR-146b−/− mice stimulated with psoriasis-associated cytokines as compared to wild type mice. Our results indicate that besides miR-146a, miR-146b is expressed and might be capable of modulation of inflammatory responses and keratinocyte proliferation in psoriatic skin.

The Autoimmune Regulator (AIRE) protein is expressed in thymic medullary epithelial cells, where it promotes the ectopic expression of tissue-restricted antigens needed for efficient negative selection of developing thymocytes. Mutations in AIRE cause APECED syndrome, which is characterized by a breakdown of self-tolerance. The molecular mechanism by which AIRE increases the expression of a variety of different genes remains unknown. Here, we studied AIRE-regulated genes using whole genome expression analysis and chromatin immunoprecipitation. We show that AIRE preferentially activates genes that are tissue-specific and characterized by low levels of initial expression in stably transfected HEK293 cell model and mouse thymic medullary epithelial cells. In addition, the AIRE-regulated genes lack active chromatin marks, such as histone H3 trimethylation (H3K4me3) and acetylation (AcH3), on their promoters. We also show that during activation by AIRE, the target genes acquire histone H3 modifications associated with transcription and RNA polymerase II. In conclusion, our data show that AIRE is able to promote ectopic gene expression from chromatin associated with histone modifications characteristic to inactive genes.

Autoimmune regulator (Aire) has a unique expression pattern in thymic medullary epithelial cells (mTECs), in which it plays a critical role in the activation of tissue-specific antigens. The expression of Aire in mTECs is activated by receptor activator of nuclear factor κB (RANK) signaling; however, the molecular mechanism behind this activation is unknown. Here, we characterize a conserved noncoding sequence 1 (CNS1) containing two NF-κB binding sites upstream of the Aire coding region. We show that CNS1-deficient mice lack thymic expression of Aire and share several features of Aire-knockout mice, including downregulation of Aire-dependent genes, impaired terminal differentiation of the mTEC population, and reduced production of thymic Treg cells. In addition, we show that CNS1 is indispensable for RANK-induced Aire expression and that CNS1 is activated by NF-κB pathway complexes containing RelA. Together, our results indicate that CNS1 is a critical link between RANK signaling, NF-κB activation, and thymic expression of Aire.Keywords: Autoimmune regulator · Enhancer · NF-κB · Receptor activator of nuclear factor κB · Thymic medulla See accompanying Commentary by Mitsuru MatsumotoAdditional supporting information may be found in the online version of this article at the publisher's web-site IntroductionCentral tolerance is achieved in the thymus by selection of a T-cell repertoire that is nonreactive to self. In the thymus, developing T cells, termed thymocytes, interact with cortical thymic epithelial cells and thymic medullary epithelial cells (cTECs and mTECs) and dendritic cells that present antigens during the positive and negative selection processes [1,2]. Due to their unique ability to express a wide range of peripheral tissue-specific antigens (TSAs), mTECs are believed to play a central role in the negative selection of self-reactive thymocytes and thereby securing tolerance to self-proteins [3,4]. The promiscuous expression of peripheral antigens in mTECs is largely controlled by autoimmune regulator Correspondence: Dr. Pärt Peterson e-mail: part.peterson@ut.ee (Aire) that constitutively promotes the transcription of hundreds of tissue-specific genes [5][6][7][8]. The importance of Aire in negative selection is exemplified by the mutations in the human Aire gene, which cause a monogenic disorder called autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy [9][10][11]. Similar to human autoimmune polyendocrinopathy-candidiasisectodermal dystrophy patients, Aire-deficient mice exhibit a multiorgan autoimmune phenotype [5], although some clear differences exist including a lack of endocrine organ involvement, relatively mild disease, and the lack of high-titer neutralizing auto-Abs to type 1 interferons and interleukin 22 [12,13].The Aire protein has a unique and highly specific expression profile. Aire is rapidly upregulated for a few days [14,15] [17,24,[26][27][28]. RANK ligation activates both the classical and alternative pathways of NF-κB signaling [29]. The classical pathway involves t...

The autoimmune regulator (AIRE) is essential for the establishment of central tolerance and prevention of autoimmunity. Interestingly, different AIRE mutations cause autoimmunity in either recessive or dominant-negative manners. Using engineered mouse models, we establish that some monoallelic mutants, including C311Y and C446G, cause breakdown of central tolerance. By using RNAseq, ATACseq, ChIPseq, and protein analyses, we dissect the underlying mechanisms for their dominancy. Specifically, we show that recessive mutations result in a lack of AIRE protein expression, while the dominant mutations in both PHD domains augment the expression of dysfunctional AIRE with altered capacity to bind chromatin and induce gene expression. Finally, we demonstrate that enhanced AIRE expression is partially due to increased chromatin accessibility of the AIRE proximal enhancer, which serves as a docking site for AIRE binding. Therefore, our data not only elucidate why some AIRE mutations are recessive while others dominant, but also identify an autoregulatory mechanism by which AIRE negatively modulates its own expression.