Macropinocytosis is the novel mechanism of endocytosis for the GPI-anchores protein uPAR.

SAHORES MACARENA; MADSEN CRIS M; CHIABRANDO GUSTAVO; BLASI FRANCESCO

Pinamar

Congreso; XLI Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular (SAIB) 2005. X Congreso de la Panamerican Association for Biochemistry and Molecular Biology (PABMB) 2005. XX Reunión Anual de la Sociedad Argentina de Neuroquí; 2005

Sociedad Argentina de Investigación en Bioquímica y Biología Molecular (SAIB)

MACROPINOCYTOSIS IS THE NOVEL MECHANISM OF ENDOCYTOSIS FOR THE GPI ANCHORED PROTEIN UPAR Sahores, M. Macarena1, 2, 3, Madsen, Chris D.1, 2, Chiabrando, Gustavo A.3, and Blasi, Francesco1, 2.1Università Vita Salute San Raffaele, Milano, Italy, 2FIRC Institute of Molecular Oncology, Milano, Italy, and 3Departamento de Bioquímica Clínica, CIBICI-CONICET, U.NC., Córdoba, Argentina. msahores@bioclin.fcq.unc.edu.ar uPAR is internalized via uPA:PAI-1 complex- clathrin- and LRP-dependent pathway. We have shown that uPAR was internalized by a constitutive mechanism in a uPA:PAI-1- clathrin- LRP-, raft-, and Rac1/RhoA/cdc42- independent fashion. Here, we used a novel protocol for flow cytometry in order to study the endocytosis of uPAR in HEK293-uPAR cells. Amiloride, a drug known to inhibit macropinocytosis, decreased the rate of internalized uPAR in a dose-dependent manner. Amiloride also inhibited the endocytosis of 70kDa-Dextran (a marker for macropinocytosis), CD59 (another GPI-anchored protein) and ATF (the amino terminal fragment of uPA that binds to uPAR). Interestingly, amiloride was unable to inhibit the endocytosis of Transferrin, LRP or uPA:PAI-1, known to be internalized via clathrin-coated pits. Thus, at least two endocytic routes for uPAR internalization might exist. One is uPA:PAI-1-induced, which is mediated by LRP and clathrin-coated pits. The second one does not require ligands, even though seems to internalize uPAR alone and uPAR-uPA complexes as well, it is constitutive and occurs via macropinocytosis, as a common mechanism for other GPI anchored proteins.