The extracellular domain of the TGFbeta type II receptor regulates membrane raft partitioning.
Cell-surface TGFbeta (transforming growth factor beta) receptors partition into membrane rafts and the caveolin-positive endocytic compartment by an unknown mechanism. In the present study, we investigated the determinant in the TGFbeta type II receptor (TbetaRII) that is necessary for membrane raft/caveolar targeting. Using subcellular fractionation and immunofluorescence microscopy techniques, we demonstrated that the extracellular domain of TbetaRII mediates receptor partitioning into raft and caveolin-positive membrane domains. Pharmacological perturbation of glycosylation using tunicamycin or the mutation of Mgat5 [mannosyl(alpha-1,6)-glycoprotein beta-1,6-N-acetylglucosaminyltransferase V] activity interfered with the raft partitioning of TbetaRII. However, this was not due to the glycosylation state of TbetaRII, as a non-glycosylated TbetaRII mutant remained enriched in membrane rafts. This suggested that other cell-surface glycoproteins associate with the extracellular domain of TbetaRII and direct their partitioning in membrane raft domains. To test this we analysed a GMCSF (granulocyte/macrophage colony-stimulating factor)-TbetaRII chimaeric receptor, which contains a glycosylated GMCSF extracellular domain fused to the transmembrane and intracellular domains of TbetaRII. This chimaeric receptor was found to be largely excluded from membrane rafts and caveolin-positive structures. Our results indicate that the extracellular domain of TbetaRII mediates receptor partitioning into membrane rafts and efficient entrance into caveolin-positive endosomes.
Samuel Lunenfeld Research Institute, Mount Sinai Hospital and the Department of Molecular and Medical Genetics, University of Toronto, Toronto, Ontario M5G1X5, Canada.
Valbona
V
Luga
Sarah
S
McLean
Christine
C
Le Roy
Maureen
M
O'Connor-McCourt
Jeffrey L
JL
Wrana
Gianni M
GM
Di Guglielmo
D000818:Animals; D051242:Caveolin 1 / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism*; D006031:Glycosylation; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D006801:Humans; D021962:Membrane Microdomains / Q000378:metabolism*; D008907:Mink; D009154:Mutation; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000378:metabolism*; D000077294:Receptor, Transforming Growth Factor-beta Type II; D018125:Receptors, Transforming Growth Factor beta / Q000737:chemistry / Q000378:metabolism*; D011994:Recombinant Proteins; D014415:Tunicamycin / Q000494:pharmacology
D051242:Caveolin 1; D018125:Receptors, Transforming Growth Factor beta; D011994:Recombinant Proteins; D014415:Tunicamycin; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D017346:Protein Serine-Threonine Kinases; D000077294:Receptor, Transforming Growth Factor-beta Type II