We recently identified collectin placenta 1 (CL-P1) from placental cDNA which has also been classified as a collectin gene (COLEC12), which is a C-type lectin containing an inner collagen-like region. CL-P1 is a type II transmembrane protein which resembles Class A scavenger receptors (SR-A) in that the scavenger receptor cysteine-rich domain is replaced by a CRD. CL-P1 is able to recognize and endocytose oxidized low density lipoprotein (OxLDL) as well as microbes. Firstly, we screened a placental cDNA library using a yeast two-hybrid system to detect molecules associated with the cytoplasmic domain of CL-P1, and identified the m2 subunit of the AP-2 adaptor complex. AP-2m2, a clathrin-associated molecule found in plasma-membrane-associated endocytic coated pits, binds to the CL-P1 N-terminal region. Analyses of the binding and endocytosis of the GalNAc sugar antigen and OxLDL by CL-P1 transfectants showed that these processes were dependent on CL-P1 expression. The CL-P1-mediated endocytosis was inhibited by tyrphostin A23 which is a specific inhibitor of tyrosine kinase, especially in m2-dependent endocytosis. However, CL-P1-mediated phagocytosis with zymosan (fungi antigen) was not inhibited by tyrphostin A23. Site-directed mutagenesis in the CL-P1 N-terminal revealed a tyrosine-based YXXf motif in its cytoplasmic region that was essential for its endocytosis of OxLDL in both HeLa and CHO transfectants. Furthermore, the endocytosis of OxLDL was disrupted in cells depleted of clathrin, adaptor AP-2 and dynamin-2. These findings indicate that tyrosine-based YXXf sequences play an important role in CL-P1-mediated OxLDL endocytosis associated with AP-2m2, and suggest that the CL-P1-ligand complexes are internalized by a clathrin-dependent and dynamin-2-dependent pathway. CL-P1 is able to endocytose OxLDL and sugar ligands for small size molecules through clathrin and dynamin-2-dependent pathway as well as to endocytose Gram-negative and Gram-positive bacteria, and yeast through another phagocytosis pathway.
