Wubbolts R, Fernandez-Borja M, Oomen L, Verwoerd D, Janssen H, Calafat J, Tulp A, Dusseljee S, Neefjes J
Department of Cellular Biochemistry, The Netherlands Cancer Institute, Amsterdam, The Netherlands.
J Cell Biol. 1996 Nov;135(3):611-22. doi: 10.1083/jcb.135.3.611.
Newly synthesized MHC class II molecules are sorted to lysosomal structures where peptide loading can occur. Beyond this point in biosynthesis, no MHC class II molecules have been detected at locations other than the cell surface. We studied this step in intracellular transport by visualizing MHC class II molecules in living cells. For this purpose we stably expressed a modified HLA-DR1 beta chain with the Green Fluorescent Protein (GFP) coupled to its cytoplasmic tail (beta-GFP) in class II-expressing Mel JuSo cells. This modification of the class II beta chain does not affect assembly, intracellular distribution, and peptide loading of the MHC class II complex. Transport of the class II/ beta-GFP chimera was studied in living cells at 37 degrees C. We visualize rapid movement of acidic class II/beta-GFP containing vesicles from lysosomal compartments to the plasma membrane and show that fusion of these vesicles with the plasma membrane occurs. Furthermore, we show that this transport route does not intersect the earlier endosomal pathway.
新合成的MHC II类分子被分选到溶酶体结构中，在那里可以发生肽加载。在生物合成的这一阶段之后，除了细胞表面外，未在其他位置检测到MHC II类分子。我们通过在活细胞中观察MHC II类分子来研究细胞内运输的这一步骤。为此，我们在表达II类分子的Mel JuSo细胞中稳定表达了一种修饰的HLA-DR1β链，其细胞质尾巴与绿色荧光蛋白（GFP）偶联（β-GFP）。II类β链的这种修饰不影响MHC II类复合物的组装、细胞内分布和肽加载。在37℃下研究了活细胞中II类/β-GFP嵌合体的运输。我们观察到含有酸性II类/β-GFP的囊泡从溶酶体区室快速移动到质膜，并表明这些囊泡与质膜发生融合。此外，我们表明这条运输途径不与早期的内体途径相交。
