Fig. 5

Relationship between CLD, the BBM and lipid metabolism enzymes studied in Caco-2 cells (a, b, c) or in mouse intestine (d). Caco-2 cells were stained with cell mask (b, c), then incubated for 10 s (a), 5 s (c) with apical mixed micelles, then fixed with EDAC and incubated with the LD540 dye to stain CLD (shown in green). a) Z-stack showing nascent CLD situated just underneath the BBM (Z = 0 μm) at upper left position to deeper in the cytosol at lower right. b) In control cells, only plasma membranes are labelled by the Cell mask TM (red staining) and intracellular lipid droplets are absent. c) After 5 s of an apical lipid load, Cellmask is present in plasma membranes (red arrows) but also coating nascent intracellular CLD (green arrows) but is not internalized in the cytosol of cells devoid of CLD (white arrows), Bar: 10 μM. d) Detection of apical biotinylated proteins in the I homogenate (Ia, Ib), in D1, D2, D3 and in M (membranes). Ia and Ib are two time chemiluminescent exposures of the same sample. Equivalent amounts of proteins were loaded and molecular weights are noted in kDalton (kDa). The experiments shown are representative of two separate experiments