engleski

Conformation based protein sorting: partitioning of empty and full MHC class I molecules in the endosomal compartments

The aim of this study was to investigate the process of sorting and localization of MHC class I molecules in functional domains of endosomal compartments and the influence of molecule conformation in that process on non-polarized cell lines. Material and methods: The kinetics of surface molecules was followed by flow cytometry and their intracellular traveling by confocal microscopy after binding of appropriate mAbs in various internalization protocols. Internalized molecules were colocalized either to each other, or with markers of endosomal compartments. In order to investigate the fate of the internalized molecules, cell surface glycoproteins were biotinylated and determined by immunoprecipitation and chemiluminiscence. Results: We have found that conformed and non-conformed MHC-I molecules did not colocalize significantly inside the cell, both at the steady state and after internalization. Soon after their endocytosis, conformed and non-conformed MHC-I molecules colocalized in the EEA1-positive early endosomes, but after that the level of their colocalization decreased. Conformed MHC-I molecules entered the recycling route, while non-conformed MHC-I molecules were mainly directed into late endosomal compartments (LEs) and colocalized with Lamp1 and LBPA. Conformed MHC-I molecules can also reach LEs, but they are partitioned from non-conformed molecules either in separated vesicles or into different subcompartments. Conclusion: Sorting of MHC-I molecules is conformation dependent. A majority of non-conformed molecules is targeted into late endosomal compartments where they are retained. Otherwise, a great deal of conformed MHC-I molecules is recycled, while a part is targeted to late endosomes, where they are mainly separated from non-conformed molecules.

MHC class I molecules

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