engleski

Down-Modulation of the Cell Surface MHC-I Molecules by MCMV Virus to Evade Immune Recognition

One strategy used by murine cytomegalovirus (MCMV) to evade immune recognition of infected cells is to down-modulate major histocompatibility complex (MHC) class I (MHC-I) expression at the plasma membrane by early viral proteins. The aim of our study was to investigate the endocytosis of MHC-I molecules in MCMV infected embryonic fibroblasts. The kinetics of MHC-I molecules internalization was followed by flow cytometry. Mechanisms of MHC-I molecules internalization were studied by using a chemical inhibitors of clathrin and caveolar endocytosis. In order to follow the internalization pathways of MHC-I molecules we used a palette of endosomal protein markers. The early endosome compartment was identified by the early endosomal marker EEA1 and by uptake of fluorescent transferrin by clathrin-dependent receptor-mediated endocytosis. The Golgi compartment was identified by the GM130 and by entry of fluorescent cholera toxin B subunit (CTB) taken up by caveolar-dependent endoytosis. Lamp-1 was used to identify late endosomes and lysosomes. The colocalization experiments were performed by immunofluorescent confocal microscopy. In order to investigate the fate of the internalized molecules, the MHC-I molecules were determined by immunoprecipitation after cell-surface biotinylation and chase period of 24 hours. The inhibitors of lysosomal and proteasomal degradation were used to distinguish the role of these two degradation pathways. In the presence of MCMV, MHC-I molecules (Kd and Dd) are almost completely down-regulated from the cell surface at 7 hours post infection. Specific inhibitors for clathrin and caveolar endocytic pathway had no effect on the MCMV-induced down-modulation of MHC-I. Thus, surface down-modulation of MHC-I molecules does not require neither clathrin-dependent nor caveolar-dependent pathway. Internalized MHC-I molecules are backsorted in endocytic compartments, trafficking through early endosomes, reaching perinuclear Golgi compartment, the place of accumulation and further tendency to be degraded. While MHC-I Kd molecules are still present after 24 hours post infection, Dd molecules are being degraded on lysosomes.

MHC-I molecules; MCMV virus; endocytotic pathway

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