Basic fibroblast growth factor (FGF-2) is addressed to caveolae after binding to the plasma membrane of BHK cells

bFGF endocytosis in BHK cells was examined by electron microscopy using a conjugate of recombinant human bFGF and digoxigenin (bFGF-DIG). This probe keeps the biological activity of non-labeled bFGF and can be readily detected with anti-digoxigenin antibodies (Gleizes et al., Anal, Biochem, 219, 360-357 (1994)). Time-course studies of bFGF-DIG endocytosis were performed by incubating BHK cells at 4 degrees C in the presence of first 20 ng/ml bFGF-DIG and then antidigoxigenin antibodies absorbed onto 10-nm gold particles. A semiquantitative study revealed that caveolae were the main endocytic pathway of bFGF-DIG in these cells, whereas clathrin-coated pits were scarcely labeled. After occurring in caveolae, bFGF-DIG was sequentially detected in tubulovesicular early endosomes, multivesicular late endosomes, and lysosomes. Under the same conditions, low density lipoprotein (LDL)-gold was seen entering the cell exclusively, through clathrin-coated pits. However, LDL-gold and bFGF-DIG were colocalized, at least in part, in common endosomal structures. Pretreatment of the cells with phosphatidylinositol-phospholipase C reduced the proportion of membrane-bound bFGF-DIG in caveolae, but did not inhibit bFGF-DIG presence in caveolae. These data suggest that bFGF enters into BHK cells through caveolae and is then shuttled into a degradative pathway similar to that of LDL.