Binding of nucleosomes to a cell surface receptor: redistribution and endocytosis in the presence of lupus antibodies.

摘要: In the present study, we sought evidence for a surface nucleosome receptor in fibroblastic cell line CV-1, and questioned whether anti-double-stranded (ds) DNA and/or anti-histone autoantibodies could recognize influence fate of surface-bound nucleosomes. 125I-labeled mononucleosomes were shown to bind layer specific, concentration-dependent saturable manner. Scatchard analysis revealed presence two binding sites: high-affinity site with Kd approximately 7nM low-affinity (Kd 400 nM) high capacity 9 x 10(7) sites. Visualization bound by fluorescence staining on both extracellular matrix (ECM). Purified mononucleosome-derived ds (180-200 bp) was found complete 125I-mononucleosomes site, stain exclusively ECM immunofluorescence, precipitate three specific proteins 43, 180 240 kDa from 125-I-labeled lysates. Nucleosomes not only 180-kDa DNA-reactive component, but also unique protein 50 kDa, suggesting that this is nucleosomes these fibroblasts. Once surface, recognized secondarily complexed lupus anti-ds or antibodies (i.e. anti-nucleosome antibodies), thus forming immune complexes situ. The complexing auto-antibodies dramatically enhance kinetics mononucleosome internalization. Following internalization nucleosome-anti-nucleosome observed formation vesicles at edge cells 5-10 min which moved toward perinuclear region 20-30 min. By means double-fluorescence labeling proteolytic treatment, fluorescent be cytoplasm, true endocytosis complexes. As confocal microscopy, no stage endocytic process there any indication coated pits participated. Co-distribution regions rich actin filaments inhibition disruption microfilament network cytochalasin D suggest mechanism mediated cytoskeleton. Taken together, our data provide receptor. We show can form situ endocytosis.