Ficolins are pattern recognition molecules that share structural similarities with mannose-binding lectin (MBL), whose key function is to recognize carbohydrates on microorganisms or epitopes on dying host cells and subsequently are capable of activating the complement system, through the lectin pathway. In humans, three ficolin genes have been identified; FCN1, FCN2 and FCN3, which encode Ficolin-1, Ficolin-2 and Ficolin-3, respectively. Apart from human, ficolins have been identified in mice, rats, chickens, pigs, hedgehogs, frogs and ascidians. However, rodents and pigs only produce two ficolins designated Ficolin-A (or -α) and Ficolin-B (or -β), which are closely related to the human FCN1 and FCN2 genes. Nevertheless, in mice a pseudogene corresponding to the human FCN3 gene has also been identified. Currently, Ficolin-3 has only been purified and characterized in human. Thus, we aimed to identify and characterize the FCN1, FCN2 and FCN3 in primates. We found that the exon organisation is very similar between all the primates included in this study and humans. Several novel inter-individual variations were found in all three FCN genes within each species. These results indicate that all the investigated primates have functional FCN1, FCN2 and FCN3 genes that are closely related to the human FCN1, FCN2 and FCN3 genes, respectively. Moreover, two polymorphisms (FCN1: Arg93Gln, FCN2: Ala185Thr) previously described in human were found in the chimpanzee and rhesus macaque, respectively. These findings indicate that the FCN genes may thus have an ancient origin. Furthermore, we showed that higher and lower primates express Ficolin-2 and Ficolin-3 in serum that has the characteristic oligomeric structures as seen in humans. This is the first time Ficolin-3 is shown outside the human situation. Serum levels of Ficolin-2 and Ficolin-3 were measured in chimpanzees and Ficolin-2 was found to be expressed in similar concentrations as found in humans. By contrast, serum Ficolin-3 was expressed in low concentrations compared to human. Finally, we showed that chimpanzee serum contains all the complement components needed to activate and generate MAC formation of the classical, the MBL and the alternative pathway of the complement system. In the human FCN genes 22 non-synonymous SNPs have been described, which were made recombinant in CHO cells and subsequently the expression level and the oligomeric structure were analysed. The variants rF1W279stop, rF2A264fs and rF3L117fs were not expressed. Normal oligomeric structures were observed for the rest of the variants. Binding studies revealed that the Ficolin-3 variants E166D and V287A showed a gain-of-function and a loss-of-function towards acetylated BSA, respectively, compared to the wild-type.
|Uddannelser||Molekylærbiologi, (Bachelor/kandidatuddannelse) Kandidat|
|Udgivelsesdato||27 maj 2010|