Naslov
Spontaneous endocytosis of MHC class I molecules on P815 cells and murine embrional fibroblasts

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

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

Izvornik
FEBS Lecture Course on Cellular Signaling & 4th Dubrovnik Signaling Conference : Abstract book ; P125 / Đikić, Ivan ; Husnjak, Koraljka - Zagreb : Institut Ruđer Bošković, 2004, 186-186

Skup
FEBS Lecture Course on Cellular Signaling ; Dubrovnik Signaling Conference (4 ; 2004)

Mjesto i datum
Dubrovnik, Hrvatska, 21.05.2004. - 27.05.2004

Vrsta sudjelovanja
Poster

Vrsta recenzije
Međunarodna recenzija

Ključne riječi
spontaneous endocytosis; induced endocytosis; MHC class I molecules

Sažetak
The aim of our study was to establish a model for spontaneous endocytosis of cell surface glycoproteins (MHC class I molecules) and to compare it with monoclonal antibody induced endocytosis on nonpolarized cell lines: nonadherent P815 cells, and adherent murine embrional fibroblasts (MEF). In order to achieve the conditions of spontaneous endocytosis on mentioned cell lines, we prevented formation of new proteins by cycloheximide (CHX), an inhibitor of protein synthesis. The kinetics of internalization of MHC class I molecules was followed by flow cytometry. In order to investigate the fate of the internalized molecules, the MHC class I molecules were determined by immunoprecipitation after cell-surface biotinylation and chase for period of 24 hours. CHX treatment inhibited exit of the new synthesized molecules to the cell surface without significant alteration of cells during the 24 hrs, and without alteration of the endocytotic process. Half-life of Kd and Dd molecules at the cell surface of P815 cells is 15-18 hrs and more than 24 hrs, respectively, and the Ld molecules are internalized from the cell surface with the half-life of about 6-10 hrs. This kinetics of the spontaneous internalization is slower than internalization kinetics after the mAb binding. It is important to note that the presence of CHX did not alter the kinetics of internalization induced by mAb binding. Fully conformed MHC class I molecules did not internalize by caveolar, nor clathrine endocytosis Incompletely conformed Ld molecules (empty molecules without bound peptide) were internalized by caveolar endocytosis on P815 cells irrespectively of the model used for induction of internalization. Furthermore, we have found that the presence of CHX only slightly influenced the kinetics of class I molecules degradation. Using the CHX we established the model of spontaneous endocytosis and we studied basic mechanism of MHC class I molecules backsorting from the cell surface. It enabled us to conclude that mechanism of their internalization depends on their stability, more than to the investigated cell type.

Izvorni jezik
Engleski

Znanstvena područja
Temeljne medicinske znanosti

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