The endocytic itineraries of lipid raft markers, such as glycosyl phosphati
dylinositol (GPI)anchored proteins and glycosphingolipids, are incompletely
understood. Here we show that different GPI-anchored proteins have differe
nt intracellular distributions; some (such as the folate receptor) accumula
te in transferrin-containing compartments, others (such as CD59 and GPI-lin
ked green fluorescent protein [GFP]) accumulate in the Golgi apparatus. Sel
ective photobleaching shows that the Golgi pool of both GPI-GFP and CD59-GF
P constantly and rapidly exchanges with the pool of these proteins found on
the plasma membrane (PM). We visualized intermediates carrying GPI-GFP fro
m the Golgi apparatus to the PM and separate structures delivering GPI-GFP
to the Golgi apparatus.
GPI-GFP does not accumulate within endocytic compartments containing transf
errin, although it is detected in intracellular structures which are endoso
mes by the criteria of accessibility to a fluid phase marker and to cholera
and shiga toxin B subunits (CTxB and STxB, which are also found in rafts).
GPI-GFP and a proportion of the total CTxB and STxB taken up into cells ar
e endocytosed independently of clathrin-associated machinery and are delive
red to the Golgi complex via indistinguishable mechanisms, Hence, they ente
r the Golgi complex in the same intermediates, get there in dependently of
both clathrin and rab5 unction, and are excluded from it at 20 degreesC and
under conditions of cholesterol sequestration. The PM-Golgi cycling pathwa
y followed by GPI-GFP could serve to regulate lipid raft distribution and f
unction within cells.