ATPase-deficient hVPS4 impairs formation of internal endosomal vesicles and stabilizes bilayered clathrin coats on endosomal vacuoles.
Epidermal growth factor receptors (EGFRs) destined for lysosomal degradation are sorted in the early endosomal vacuole into small, lumenal vesicles that arise by inward budding of the limiting membrane. We have previously shown that, before their incorporation into internal vesicles, EGFRs are concentrated in flat bilayered-clathrin coats on the endosomal vacuole. Here, we show that an ATPase-deficient mutant of hVPS4 (hVPS4(EQ)) increases the association of bilayered coats with endosomal vacuoles. In addition, hVPS4(EQ) leads to a reduction in the number of internal vesicles in early and late endosomal vacuoles, and retention of EGFRs at the limiting membrane. Interestingly, hVPS4(EQ) was predominantly found on non-coated regions of endosomal vacuoles, often at the rim of a coated area. In line with published data on Vps4p function in yeast, these results suggest that hVPS4 is involved in the release of components of the bilayered coat from the endosomal membrane. Moreover, our data suggest that disassembly of the coat is required for the formation of internal vesicles.
Sachse
Martin
M
Department of Cell Biology, University Medical Center and Institute for Biomembranes, 3584 CX Utrecht, The Netherlands.
Strous
Ger J
GJ
Klumperman
Judith
J
D000074183:ATPases Associated with Diverse Cellular Activities; D000251:Adenosine Triphosphatases / Q000172:deficiency* / Q000235:genetics / Q000378:metabolism*; D022163:Clathrin-Coated Vesicles / Q000378:metabolism* / Q000648:ultrastructure; D056827:Endosomal Sorting Complexes Required for Transport; D011992:Endosomes / Q000378:metabolism* / Q000648:ultrastructure; D066246:ErbB Receptors / Q000378:metabolism; D015870:Gene Expression; D006367:HeLa Cells; D006801:Humans; D009154:Mutation; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D025262:Vacuolar Proton-Translocating ATPases; D014617:Vacuoles / Q000378:metabolism* / Q000648:ultrastructure; D033921:Vesicular Transport Proteins
D056827:Endosomal Sorting Complexes Required for Transport; D012097:Repressor Proteins; D033921:Vesicular Transport Proteins; D066246:ErbB Receptors; D000251:Adenosine Triphosphatases; D025262:Vacuolar Proton-Translocating ATPases; D000074183:ATPases Associated with Diverse Cellular Activities; C490305:VPS4A protein, human