טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
Ph.D Thesis
Ph.D StudentNatalie Dror
SubjectCharacterization of IRF-8 and IRF-1 Regulatory Network in
Mature Macrophages
DepartmentDepartment of Biotechnology and Food Engineering
Supervisor Full Professor Levi Ben-Zion
Full Thesis textFull thesis text - English Version


Abstract

IRF1 and IRF8 (Interferon Refulatory Factors 1 & 8) are important regulators of macrophage differentiation and function. These factors exert some of their activities through the formation of heterocomplexes. As such, they are coactivators of various Interferon (IFN)-inducible genes in macrophages. To gain better insights into the involvement of these two transcription factors in innate immunity and to identify their regulatory network in activated macrophages, we employed DNA microarray analysis.  We looked for changes in the transcriptome profile of peritoneal macrophages from wild type mice and from null mice to IRF1 and IRF8 before and following 4hrs exposure to IFN-γ and LPS. The expression level of  265 transcripts did not change in both KO strains following macrophage activation. Out of these, ten genes were selected for additional study due to their known functions in processes involved in innate immunity. The role of IRF1 and IRF8 in their regulated expression in myeloid cells was further confirmed by Real-Time PCR and by promoter analyses. Last, we focused on the Promyelocytic Leukemia (PML) gene. PML acts as a growth and tumor suppressor , cell cycle regulator, mediator of apoptosis, and regulator of transcription, translation, and hematopoietic differentiation. PML is concentrated in Nuclear Bodies, whose function is not entirely understood. IFNs induce PML’s expression through specific promoter motifs, the ISRE and GAS elements, that are IFN-responsive. Our results clearly demonstrated that IRF8 is an obligatory regulator of PML in macrophages, leading to the expression of the PML-I isoform.  IRF8 is essential for the induced expression of PML in activated macrophages and for the formation of NBs. Reduced IRF8 transcript levels were reported in CML patients and recovery to normal levels were observed in remised patients treated with IFN-α. We observed a significant correlation between the levels of IRF8 and PML in these CML patients. Together, our results suggest a broader role for IRF1 and IRF8 in macrophages differentiation and maturation, being important mediators of innate immunity. Furthermore, the great significance that emerges from our results points to the important role of IRF8 in monopoiesis as well as in myeloleukemia suppression, through the regulation of PML. When IRF8 levels are compromised, the reduced PML expression may lead to genome instability and eventually to the leukemic phenotype.