Semen clusterin targets extracellular proteins to dendritic cells for antigen presentation

MERLOTTI IPPÓLITO, ANTONELLA; DANTAS, EZEQUIEL; DUETTE, GABRIEL; ERNST, GLENDA; PEREYRA, PEHUÉN; REMES LENICOV, FEDERICO; VARESE, AUGUSTO; GEFFNER, JORGE; SABATTÉ, JUAN

Los Cocos

Congreso; LXI Reunión Anual de la Sociedad Argentina de Inmunología; 2013

Sociedad Argentina de Inmunología

Clusterin is an ubicuos glycoprotein present in different tissues and fluids. One of the properties of serum clusterin (S-CLU) is its ability to interact with unfolded proteins avoiding their aggregation, acting as an extracellular chaperone. In contrast to serum clusterin, seminal plasma form (PS-CLU) presents a glycosilation pattern with high fucose arrangements, which allows it bind with high affinity to the pattern recognition receptor DC-SIGN expressed on DCs. The aim of this work is to analyze if PS-CLU could interact with stressed proteins and target them to DC-SIGN on dendritic cells (DCs) promoting antigen endocytosis and presentation. We purified clusterin from seminal plasma of healthy donors wtih affinity chromatography. Dendritics cells were differentiated from monocytes of peripheral blood by culturing them during 5 days with GM-CSF (40ng/ml) and IL-4 (20ng/ml). Stressing BSA (1mg/ml) in presence or absence of PS-CLU (1mg/ml) during 30 min at 60°C, and reading the solution turbidity at 360 nm as a measured of stressed protein, we observed that PS-CLU inhibits protein aggregation (BSA: 0.545 vs. BSA-CLU: 0.270). We saw by flow citometry in binding assays to Raji DC-SIGN positive cells a higer MFI when biotynilated BSA is stressed in CLU-PS presence (MFI BSA: 407 vs. BSA-CLU: 1695). Using DCs we observed at different times by confocal microscopy intracellular colocalization of stressed biotynilated BSA with PS-CLU, DC-SIGN and the endosomal markers EEA1 and Lamp-1. Finally, we analyzed by ELISPOT the presentation of DCs internalized antigens to T lymphocytes in presence or absence of PS-CLU. We used M. Tuberculosis derived proteins as antigen. PPD: 5 vs PPD-CLU: 20. These results suggest that fucosylated clusterin, like PS-CLU, can target stressed proteins to DC-SIGN expressed in DCs to be processed and presented by this cells.