Endocytosis of GPI-linked membrane folate receptor-alpha

被引:168
作者
Rijnboutt, S
Jansen, G
Posthuma, G
Hynes, JB
Schornagel, JH
Strous, GJ
机构
[1] UNIV UTRECHT, FAC MED, DEPT CELL BIOL, 3584 CX UTRECHT, NETHERLANDS
[2] UNIV UTRECHT, BIOMEMBRANE INST, UTRECHT, NETHERLANDS
[3] FREE UNIV AMSTERDAM HOSP, DEPT ONCOL, AMSTERDAM, NETHERLANDS
[4] NETHERLANDS CANC INST, DEPT INTERNAL MED, AMSTERDAM, NETHERLANDS
[5] MED UNIV S CAROLINA, DEPT PHARMACEUT SCI, CHARLESTON, SC 29425 USA
关键词
D O I
10.1083/jcb.132.1.35
中图分类号
Q2 [细胞生物学];
学科分类号
071009 ; 090102 ;
摘要
GPI-linked membrane folate receptors (MFRs) have been implicated in the receptor-mediated uptake of reduced folate cofactors and folate-based chemotherapeutic drugs, We have studied the biosynthetic transport to and internalization of MFR isoform alpha in KB-cells, MFR-alpha was synthesized as a 32-kD protein and converted in a maturely glycosylated 36-38-kD protein 1 h after synthesis. 32-kD MFR-alpha was completely soluble in Triton X-100 at 0 degrees C. In contrast, only 33% of the 36-38-kD species could be solubilized at these conditions whereas complete solubilization was obtained in Triton X-100 at 37 degrees C or in the presence of saponin at 0 degrees C. Similar solubilization characteristics were found when MFR-alpha at the plasma membrane was labeled with a crosslinkable I-125-labeled photoaffinity-analog of folic acid as a ligand, Triton X-100-insoluble membrane domains containing MFR-alpha could be separated from soluble MFR-alpha on sucrose flotation gradients. Only Triton X-100 soluble MFR-alpha was internalized from the plasma membrane, The reduced-folate-carrier, an integral membrane protein capable of translocating (anti-)folates across membranes, was completely excluded from the Triton X-100-resistant membrane domains, Internalized MFR-alpha recycled slowly to the cell surface during which it remained soluble in Triton X-100 at 0 degrees C. Using immunoelectron microscopy, we found MFR-alpha: along the entire endocytic pathway: in clathrin-coated buds and vesicles, and in small and large endosomal vacuoles, In conclusion, our data indicate that a large fraction, if not all, of internalizing MFR-alpha bypasses caveolae.
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页码:35 / 47
页数:13
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