İnflamatuvar Hücre Göçü Sürecinde Pyrin ve Aktin İle Ilişkili Proteinlerin Moleküler Analizi

Abstract

MEFV gene, encodes Pyrin protein, cause familial Mediterranean fever (FMF). Pyrin which is primarily expressed in neutrophils and monocytes, appears to be a regulator of inflammation, but its exact role on FMF pathophysiology is still controversial. Several studies have been demonstrated a close relation between Pyrin and the cytosekeletal elements. Therefore we hypothesized that Pyrin can play role in inflammatory cell migration. In this thesis, we analyzed known actin-related proteins that have roles in cell migration; PSTPIP2, LSP1, LPXN, CORO2A, CTNNB1, WDR1, FMNL1, DAAM1, ACTR2 and the interaction of Pyrin with these proteins during inflammatory cell migration process. Related genes were investigated on both RNA and protein levels using qRT-PCR, PCR and IF staining techniques in neutrophil-like HL-60 cells. We demonstrated that actin-related genes are expressed in HL-60 cells and RNA levels increase significantly during differentiation. MEFV expression is also increased in differentiated and differentiated-stimulated HL-60 cells. IF stainings showed that Pyrin is cytoplasmic in normal and differentiated cells but it localizes much more in polarized edge of the cells and in cell protrusions during cell migration. We also demonstrated that actin-related proteins co-localize with Pyrin in normal, differentiated and differentiated-stimulated cells. During migration, co-localization was observed mostly in polarized edges and protrusions. PSTPIP2, LPXN, LSP1 and FMNL1 proteins demonstrate a strong co-localization with Pyrin especially in migration process. This study has importance in terms of showing the potential involvement of Pyrin to actin machinery via actin-related proteins that have strong co-localization, and supportting the idea of the possible role of Pyrin in inflammatory cell migration through actin skeleton.