Detergent-insoluble GPI-anchored proteins are apically sorted in Fischer rat thyroid cells, but interference with cholesterol or sphingolipids differentially affects detergent insolubility and apical sorting
C. Lipardi et al., Detergent-insoluble GPI-anchored proteins are apically sorted in Fischer rat thyroid cells, but interference with cholesterol or sphingolipids differentially affects detergent insolubility and apical sorting, MOL BIOL CE, 11(2), 2000, pp. 531-542
In contrast to Madin-Darby canine kidney cells, Fischer rat thyroid cells d
eliver the majority of endogenous,glycosylphosphatidyl inositol (GPI)-ancho
red proteins to the basolateral surface. However, we report here that the G
PI proteins Placental Alkaline Phosphatase (PLAP) and Neurotrophin Receptor
-Placental Alkaline Phosphatase (NTR-PLAP) are apically localized in transf
ected Fischer rat thyroid cells. In agreement with the "raft hypothesis," w
hich postulates the incorporation of GPI proteins into,glycosphingolipids a
nd cholesterol-enriched rafts, we found that both of these proteins were in
soluble in Triton X-100 and floated into the lighter fractions of sucrose d
ensity gradients. However, disruption of lipid rafts by removal of choleste
rol did not cause surface missorting of FLAP and NTR-LAP, and the altered.
surface sorting of these proteins after Fumonisin B1 treatment did not corr
elate with reduced levels in Triton X-100 -insoluble fractions. Furthermore
, in contrast to the GPI-anchored forms of both of these proteins, the secr
etory and transmembrane forms (in the absence of a basolateral cytoplasmic
signal) were sorted to the apical surface without association with lipid mi
crodomains. Together, these data demonstrate that the GPI anchor is require
d to mediate raft association but is not sufficient to determine apical sor
ting. They also suggest that signals present in the ectodomain of the prote
ins play a major role and that lipid rafts may facilitate the recognition o
f these signals in the trans-Golgi network, even though they are not requir
ed for epical sorting.