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Identifier

000434026

Title

Ο ρόλος των απομεθυλασών των ιστονών, PHF8 και PHF2, στην επιγενετική ρύθμιση της ενεργοποίησης των μακροφάγων

Alternative Title

The role of histone demethylases, PHF8 and PHF2, in the epigenetic regulation of activated macrophages

Author

Αξαρλής, Κωνσταντίνος Ν.

Thesis advisor

Τσατσάνης, Χρήστος

Reviewer

Μπέρτσιας, Γεώργιος
Χαμηλός, Γεώργιος

Abstract

Macrophages are the central mediators of inflammatory responses, they constitute the basis of innate immunity, while, at the same time, exhibit an important role in several functions of homeostasis1,2 . Macrophage activation requires strict and multiple layers of gene regulation, which may lead in a vast spectrum of pro- or anti- inflammaroty phenotypes, termed as “M1” or “M2”, respectively2,3 . A growing body of evidence indicates that this enormous macrophage plasticity is excessively regulated at the epigenetic level and that different metabolic adaptations influence the outcome of the immune response. In the present postgraduate thesis, the role of histone demethylases “PHF8” (KDM7B) and “PHF2” (KDM7C) in macrophage activation was investigated, including the expression of genes and secretion of cytokines that are implicated in TLR4-induced inflammation, the impact on immunometabolic regulation, as well as macrophage phagocytosis capacity. Finally, a possible interaction of PHF8 with AKT kinases, which have a crusial role in immune response, was tested. For that purpose, stable macrophage cell-lines which under- or over- express the aforementioned proteins were created with CRISPR. Several molecular- and cell- biology techniques were used, such as si RNA on primary macrophages, FACS, RT-qPCR, ELISA, and WB. It was exhibited that PHF8 plays both a pro- and an anti- inflammatory role in macrophage activation. Furthermore, it was demonstrated that PHF8 downregulates mitochondrial biogenesis, and, by extension, oxidative phosphorylation. Overexpression of PHF8 led to elevated phatocytosis capacity. Initial indications suggested that AKT1 and AKT2 kinases control PHF8 expression in an opposite manner and that they may affect its total and phosphorylated protein levels. Histone demethylase PHF2 was found to exhibit a strong pro-inflammatory activity. It was displayed that it regulates a number of immune regulation genes in an opposite way than PHF8, suggesting a potential competitive relashionship between the two proteins. Moreover, in some cases similar gene regulation was also observed, indicating a synergistic mechanism of action. Finally, it was displayed that PHF2 is necessary for effective phagocytosis.

Language

Greek

Subject

Immunometabolism

Macrophages

TLR4-Mediated immune response

TLR4-Διαμεσολαβούμενη ανοσολογική απόκριση

Ανοσομεταβολισμός

Μακροφάγα

Issue date

2020-11-27

Collection