Autoimmune Regulator: Characterization of Thymic Gene Regulation and Promoter Methylation
Date
2012-05-15
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Abstract
Tüümuse medullaarsed epiteelirakud ekspresseerivad AIRE (Autoimmuunne Regulaator) valku, mis suunab perifeersete koespetsiifiliste geenide avatud geeniekspressiooni. Mutatsioonid AIRE geenis toovad endaga kaasa haruldase autoimmuunhaiguse APECED (Autoimmune Polyendocrinopathy Candidiasis Ectodermal Dystrophy), millega kaasneb immunoloogilise tolerantsuse kadumine mitmete endokriinorganite suhtes. Käesolevas töös leidsime, et koespetsiifiliste geenide ekspressioon sõltub Aire geeni koopiate arvust tüümuses ja Aire kui ka koespetsiifilised geenid ekspresseeruvad tüümuse medullas ning neil on sarnane avaldumismuster tüümuse erinevatel arenguetappidel. Aire geeni lisamise tingimustes tõusis koespetsiifiliste geenide mRNA tase. Tüümuse struktuuri lõhkumise tulemusena langes kõigi uuritud geenide ekpressioon. Antud tulemused näitavad, et Aire geenil on otsene roll koespetsiifiliste geenide ekspressioonil ja läbi Aire geeni on võimalik mõjutada tsentraalse tolerantsuse või autoimmuunsuse tekkimist.
Kuna tüümuse kemokiinid on olulised tüümusesse sisenevate tümotsüütide migratsioonil, siis uurisime Aire geeni rolli tüümuse kemokiinide tootmises. Leidsime, et tüümuses Aire geeni puudumise tulemusena langevad CCR4 ja CCR7 ligandide tasemed ning CCR4 ligandid on Aire poolt reguleeritud CD80hi mTEC rakkudes. Sarnaselt koespetsiifilistele antigeenidele jälgisid ka tüümuse kemokiinid Aire avaldumismustrit sünnijärgsel perioodil. Aire geeni üleekspresseerimise tingimustes tõusis CCL5, CCL22 and CCL19 kemokiinide tase. Antud leidude puhul on tegemist Aire uudse rolliga tsentraalse tolerantsuse kujunemises.
Edasi uurisime AIRE promooteri metülatsiooni ja leidsime, et AIRE promooter oli hüpometüleeritud mTECides ja cTECides ning CpG metülatsiooni tase tõusis AIRE negatiivsetes tümotsüütides. Samas oli AIRE promooter hüpometüleeritud Aire negatiivsetes tüümuse kasvajates ja perifeersetes kudedes. Ning leidsime positiivse korrelatsiooni AIRE ekspressiooni ja AIRE promooteril asuva aktiivse kromatiini märgise H3K4me3 vahel.
Thymic medullary epithelial cells express the AIRE (Autoimmune Regulator) gene, which directs the ectopic expression of peripheral tissue-specific antigens (TSAs). Mutations in the AIRE gene cause a rare autoimmune disease APECED (Autoimmune Polyendocrinopathy Candidiasis Ectodermal Dystrophy), accompanied by a loss of immunological tolerance to a number of endocrine organs. We found that TSA expression is dependent on Aire gene copy number in thymus. Aire as well as TSAs were expressed in thymic medulla and had similar expression pattern in the thymus at the different stages of development. TSA expression increased in the conditions of Aire over-expression. The destruction of the thymus structure resulted in a drop of expression of TSAs. These results suggest that the Aire gene has a direct role in TSA expression and via Aire gene it should be possible to modulate the negative selection of autoreactive T cells. Since thymic chemokines are important for the migration of thymocytes into the thymus, we examined the role of the Aire gene on thymic chemokine production. We found that the thymic absence of Aire gene resulted in decreased levels of CCR4 and CCR7 ligands, whereas CCR4 ligands were regulated by Aire in CD80-positive thymic medullary epithelial cells. Similarly to TSAs in the thymus, the chemokines follow the Aire expression pattern after birth. In over-expression of the Aire gene, the expression of CCL5, CCL22 and CCL19 chemokines was also increased. We describe a novel role for Aire in the development of central tolerance. Furthermore, during the studies of genomic methylation signatures in Aire promoter region in different cells and tissues, we found that the AIRE promoter was hypomethylated in thymic epithelial cells but highly methylated in thymocytes. Also, the AIRE promoter was hypomethylated in AIRE-negative thymic epithelial tumors (thymomas) and in several peripheral tissues. Positive correlation was found in human thymic epithelial cells between AIRE expression and an active chromatin mark; the histone H3K4me3 modification at the AIRE promoter.
Thymic medullary epithelial cells express the AIRE (Autoimmune Regulator) gene, which directs the ectopic expression of peripheral tissue-specific antigens (TSAs). Mutations in the AIRE gene cause a rare autoimmune disease APECED (Autoimmune Polyendocrinopathy Candidiasis Ectodermal Dystrophy), accompanied by a loss of immunological tolerance to a number of endocrine organs. We found that TSA expression is dependent on Aire gene copy number in thymus. Aire as well as TSAs were expressed in thymic medulla and had similar expression pattern in the thymus at the different stages of development. TSA expression increased in the conditions of Aire over-expression. The destruction of the thymus structure resulted in a drop of expression of TSAs. These results suggest that the Aire gene has a direct role in TSA expression and via Aire gene it should be possible to modulate the negative selection of autoreactive T cells. Since thymic chemokines are important for the migration of thymocytes into the thymus, we examined the role of the Aire gene on thymic chemokine production. We found that the thymic absence of Aire gene resulted in decreased levels of CCR4 and CCR7 ligands, whereas CCR4 ligands were regulated by Aire in CD80-positive thymic medullary epithelial cells. Similarly to TSAs in the thymus, the chemokines follow the Aire expression pattern after birth. In over-expression of the Aire gene, the expression of CCL5, CCL22 and CCL19 chemokines was also increased. We describe a novel role for Aire in the development of central tolerance. Furthermore, during the studies of genomic methylation signatures in Aire promoter region in different cells and tissues, we found that the AIRE promoter was hypomethylated in thymic epithelial cells but highly methylated in thymocytes. Also, the AIRE promoter was hypomethylated in AIRE-negative thymic epithelial tumors (thymomas) and in several peripheral tissues. Positive correlation was found in human thymic epithelial cells between AIRE expression and an active chromatin mark; the histone H3K4me3 modification at the AIRE promoter.
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Keywords
geenid, autoimmuunsus, geeniregulatsioon, kemokiinid, harkelund, genes, autoimmunity, gene regulation, chemokines, thymes