RESUMO
Spleen-resident dendritic cell (DC) populations occupy sentinel positions for the capture and presentation of blood-borne antigens. Here we found a difference in expression of the chemotactic receptor EBI2 (GPR183) on splenic DC subsets and that EBI2 regulated the positioning and homeostasis of DCs in the spleen. EBI2 and its main ligand, 7α,25-OHC, were required for the generation of the splenic CD4(+) DC subset and the localization of DCs in bridging channels. Absence of EBI2 from DCs resulted in defects in both the activation of CD4(+) T cells and the induction of antibody responses. Regulated expression of EBI2 on DC populations is therefore critical for the generation and correct positioning of splenic DCs and the initiation of immune responses.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Quimiotaxia/imunologia , Células Dendríticas/imunologia , Ativação Linfocitária , Receptores Acoplados a Proteínas G/metabolismo , Baço/imunologia , Animais , Formação de Anticorpos/genética , Apresentação de Antígeno/genética , Antígenos CD4/metabolismo , Movimento Celular/genética , Movimento Celular/imunologia , Células Cultivadas , Células Dendríticas/patologia , Homeostase/genética , Homeostase/imunologia , Ativação Linfocitária/genética , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Receptores Acoplados a Proteínas G/genética , Baço/patologiaRESUMO
Clusterin is the first well characterized, constitutively secreted extracellular chaperone that binds to exposed regions of hydrophobicity on non-native proteins. It may help control the folding state of extracellular proteins by targeting them for receptor-mediated endocytosis and intracellular lysosomal degradation. A notable feature of secreted clusterin is its heavy glycosylation. Although carbohydrate comprises approximately 20-25% of the total mass of the mature molecule, its function is unknown. Results from the current study demonstrate that deglycosylation of human serum clusterin had little effect on its overall secondary structure content but produced a small increase in solvent-exposed hydrophobicity and enhanced the propensity of the molecule to aggregate in solution. These changes were associated with increased binding to a variety of ligands but did not substantially impact the ability of clusterin to inhibit heat-induced precipitation of citrate synthase. Evidence suggesting that the normally conjugated sugars are important in the interaction of secreted clusterin with a lectin-type receptor on liver cells is also presented. Bulk expression of fully processed, glycosylated clusterin in mammalian cells is difficult, often producing inappropriately disulfide-bonded high molecular weight aggregates; this has hampered previous studies aimed at identifying those regions of the molecule important in its chaperone action. The current results suggest that it may be possible in the future to study the structure and chaperone function of clusterin using recombinant protein (lacking sugars) conveniently bulk-expressed in bacteria.
