Biochemical and functional characterization of ubiquitin-like proteines ISG15 and FAT10 in human dendritic cells.

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During the process of maturation of dendritic cells (DCs), which are essential for an effective immune response, DCs undergo fundamental changes both in gene expression, cellular function and cell mor-phology. Post-translational modifications of proteins by either ubiquitin and or ubiquitin-like proteins (Ubls) seem to be an important regulatory principle in the adaptation of cells to changing physiolo-gical requirements. Conjugation of ubiquitin-like proteins often influences processes connected with the adaptation of cells to changing physiological requirements. Two cytokine induced ubiquitin-like proteins whose functions seem to be closely related to the immune response are ISG15 and FAT10. The enzymes responsible for ISG15 conjugation are known. For FAT10, only the E1 activating enzyme has been identified. Furthermore, the functional impacts of both ISG15 and FAT10 conjugation still remain to be elucidated. Based on DC gene expression analysis we aim to identify the FAT10 E2 and E3 enzymes and study FAT10 conjugate formation in DCs both in vitro and in vivo. siRNA experi-ments will be performed to study the effects of FAT10 and ISG15 conjugation on gene and protein expression during DC maturation. In addition the intracellular pathways of ISG15 will be followed by immuno- cytological and life imaging experiments.