The influence of AIRE on gene expression – studies of transcriptional regulatory mechanisms in cell culture systems
Date
2017-11-14
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Abstract
Probleemi kirjaldus. Immuunsüsteemi põhiline ülesanne on võidelda haigustekitajatega, mis üritavad tungida organismi, et seda kahjustada. Samaväärselt oluline on immuunsüsteemi võime hoiduda kehaomaste või väljastpoolt tulevate kahjutute molekulide vastu suunatud immuunreaktsioonist. Seda nähtust nimetatakse immuuntolerantsuseks. Immuuntolerantsuse kujunemisel on suur roll harknäärmel, mis vastutab kehaomaste valkude suhtes tundlike T-rakkude kõrvaldamise eest. Suuresti sõltub see protsess ühest valgust nimega autoimmuunsuse regulaator (AIRE). AIRE olemasolul avalduvad harknäärmes kehaomased valgud, mis viiakse kokku arenevate T-rakkudega. T-rakk, millel tekib tugev reaktsioon nende valkude suhtes, sureb enne, kui jõuab harknäärmest lahkuda. Kui harknäärmes puudub AIRE, jäävad ohtlikud T-rakud ellu, siirduvad vere- ning lümfiringe kaudu teistesse kudedesse ja vallandavad autoimmuunrünnaku, mis hävitab koe, põhjustades seeläbi raskeid haiguseid.
Tulemus ja kasutegur. Käesolevas doktoritöös uurisime, kuidas mõjutab AIRE funktsiooni see, kui valgule lisada biokeemilisel viisil atsetüülrühmasid. Leidsime, et muutuvad AIRE paiknemine rakutuumas, valgu stabiilsus ning võime aktiveerida kehaomaste valkude avaldumist. Lisaks analüüsisime AIRE valgus esinevat mutatsiooni, mis põhjustab inimestel AIRE-puudulikkusest tingitud autoimmuunhaigust. Selgus, et mutatsioon lõhub AIRE valgu struktuuri ning kuigi AIRE on rakus olemas, ei pääse ta rakutuuma, et kehaomaste valkude avaldumist aktiveerida. Doktoritöö viimases osas keskendusime muutustele genoomi struktuuris, mis kaasnevad AIRE poolt aktiveeritud kehaomaste valkude avaldumisega. Avastasime vastupidiselt ootustele, et muutused toimuvad kehaomaste valkude geenidest kaugel olevates genoomiosades, viidates sellele, et AIRE-l on seni teadaolevast palju laialdasem mõju genoomi ülesehitusele.
Kokkuvõtvalt, uurides AIRE valku mõjutavaid tegureid ning AIRE enda toimet genoomi struktuurile, suudame paremini mõista immuuntolerantsuse ning autoimmuunhaiguste tekkemehhanisme.
Description of the problem. The main function of the immune system is to fight off pathogens that try to invade and harm the body. At the same time, the immune system has to block any immune reactions against harmless antigens stemming from the organism itself or from the environment. This phenomenon is called immune tolerance. The thymus plays a major role in establishing tolerance towards self-antigens by eliminating autoreactive T-cells. This process is primarily controlled by a single protein called autoimmune regulator (AIRE). AIRE promotes the expression of self-antigens that are presented to developing T-cells in the thymus. T-cell that strongly react to the self-antigens, will die before they leave the thymus. If AIRE is absent from the thymus, then the self-reactive T-cells will survive and migrate to other tissues, which can be targeted and destroyed by these T-cells causing an autoimmune disease. Result and benefit. In this thesis, we investigated the effect of acetylation of the AIRE protein. We found that it alters the localisation and stability of the protein and eventually affects the expression of self-antigens. Additionally, we analysed a mutation in the AIRE protein that causes AIRE-deficiency and autoimmunity in humans. We conclude that the mutation destroys AIRE protein structure, and although AIRE is still present in the cells, it cannot move into the nucleus to activate the expression of self-antigens. In the final part of the thesis we explored the changes in the structure of the genome that coincide with the AIRE-dependent activation of self-antigen expression. Contrary to expectations, the genomic alterations occurred far away from self-antigen coding genes suggesting that AIRE has a much broader impact on the gene regulatory processes in the nucleus than previously anticipated. In summary, uncovering factors that affect AIRE function and how AIRE itself contributes to the genomic organisation expand our understanding of the molecular mechanisms behind immune tolerance and autoimmunity.
Description of the problem. The main function of the immune system is to fight off pathogens that try to invade and harm the body. At the same time, the immune system has to block any immune reactions against harmless antigens stemming from the organism itself or from the environment. This phenomenon is called immune tolerance. The thymus plays a major role in establishing tolerance towards self-antigens by eliminating autoreactive T-cells. This process is primarily controlled by a single protein called autoimmune regulator (AIRE). AIRE promotes the expression of self-antigens that are presented to developing T-cells in the thymus. T-cell that strongly react to the self-antigens, will die before they leave the thymus. If AIRE is absent from the thymus, then the self-reactive T-cells will survive and migrate to other tissues, which can be targeted and destroyed by these T-cells causing an autoimmune disease. Result and benefit. In this thesis, we investigated the effect of acetylation of the AIRE protein. We found that it alters the localisation and stability of the protein and eventually affects the expression of self-antigens. Additionally, we analysed a mutation in the AIRE protein that causes AIRE-deficiency and autoimmunity in humans. We conclude that the mutation destroys AIRE protein structure, and although AIRE is still present in the cells, it cannot move into the nucleus to activate the expression of self-antigens. In the final part of the thesis we explored the changes in the structure of the genome that coincide with the AIRE-dependent activation of self-antigen expression. Contrary to expectations, the genomic alterations occurred far away from self-antigen coding genes suggesting that AIRE has a much broader impact on the gene regulatory processes in the nucleus than previously anticipated. In summary, uncovering factors that affect AIRE function and how AIRE itself contributes to the genomic organisation expand our understanding of the molecular mechanisms behind immune tolerance and autoimmunity.
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Keywords
koekultuurid, autoimmuunsus, transkriptsioonifaktorid, geeniekspressioon, transkriptsioon (biol.), regulatsioon (biol.), kromatiin, tissue cultures, autoimmunity, transcription factors, gene expression, transcription (biol.), regulation (biol.), chromatin