Naslov
MHC Class I molecules are recycled after their internalization, but their pathways are conformation dependent

Autori
Mahmutefendić, Hana ; Blagojević, Gordana ; Kučić, Natalia ; Lučin, Pero

Vrsta, podvrsta i kategorija rada
Sažeci sa skupova, sažetak, znanstveni

Izvornik
1st Joint Meeting of European National Societies of Immunology and 16th European Congress of Immunology, Book of abstracts / European Federation of Immunological Societies - Pariz, 2006

Skup
1st Joint Meeting of European National Societies of Immunology and 16th European Congress of Immunology

Mjesto i datum
Pariz, Francuska, 06.09.2006

Vrsta sudjelovanja
Poster

Vrsta recenzije
Nije recenziran

Ključne riječi
endocytosis; MHC-I molecules; vesicular transport

Sažetak
Problem: The aim of our study was to compare the internalization mechanisms and endocytic trafficking of conformed and non-conformed murine MHC class I (MHC-I) molecules (Kd, Dd and Ld) on non-polarized cell lines P815 mastocytoma cell line and Balb 3T3 fibroblast cell line). Material and methods: Localization of MHC class I molecules in detergent insoluble membrane domains was determined by short ice-cold Triton X treatment and confocal microscopy or immunoprecipitation. Endocytosis and intracellular trafficking of MHC-I molecules were followed in the model of spontaneous internalization after cycloheximide treatment, or after surface binding of monoclonal antibodies (mAbs). Endocytic pathway was dissected by treatment with inhibitors of endocytosis and endosomal acidification. Transformation of MHC class I molecules into misfolded conformation was achieved by short acidification of cell culture medium. Cell surface expression of MHC-I molecules was determined by flow cytometry and intracellular expression by confocal microscopy. Cell surface biotinylation, followed by immunoprecipitation and SDS-PAGE, was used to determine the kinetics of degradation. Results: Non-conformed Dd and Ld molecules were localized in the lipid rafts, and were internalized by cholesterol dependent mechanism that is not caveolar. In contrast, conformed class I molecules were internalized by the bulk pathway and did not localize in the lipid rafts. Furthermore, conformed MHC-I molecules were identified in the early recycling pathway, whereas non-conformed Ld are directed to an endosomal retention compartment (empty Ld compartment - ELC), a place where empty Ld molecules normally accumulate. After treatment with the ihibitors of endosomal acidification, conformed and non-conformed Ld molecules colocalize to the large extent, whereas in untreated cells very little colocalization can be found. After internalization, both conformed and non-conformed molecules were found in EEA1+ and Tf+ compartments, but not before 10-20 minutes after internalization. Finally, both types of class I molecules are degraded in lysosomes, but with different kinetics. Conclusion: Conformed and non-conformed MHC-I molecules are internalized by different mechanisms, but their endocytic pathways are intersected in sorting endosomes and lysosomes. Empty Ld molecules are retained in ELC and probably recycled back to the cell surface through an peptide loading compartment, or directed into lysosomes for degradation.

Izvorni jezik
Engleski

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