Annexin I is phosphorylated in the multivesicular body during the processing of the epidermal growth factor receptor.
We have previously shown that an active epidermal growth factor receptor (EGF-R) kinase is necessary for efficient sorting of the EGF-R to the lysosome, and we have shown that this occurs in the multivesicular body (MVB), where EGF-R are sorted away from recycling receptors by being removed to the internal vesicles of the MVB. The aim of the present study was to identify substrates of the EGF-R kinase associated with MVBs which might play a role in this sorting process. We used a density shift technique to isolate MVBs and show that the major substrates phosphorylated in vitro within MVBs which contain an active EGF-R kinase are the EGF-R itself and annexin I. Annexin I is associated with both plasma membrane and MVBs in a calcium-independent manner but can be phosphorylated in vitro only in MVBs. Phosphorylation of calcium-independent annexin I in isolated MVBs converts it to a form that requires calcium for membrane association. In cells with an active EGF-R kinase the amount of calcium-independent annexin I in MVBs is reduced, suggesting that a phosphorylation-induced conversion of the calcium independent to the calcium-dependent form also occurs in vivo. Our observations, together with the known properties of annexin I in mediating membrane fusion, suggest that inward vesiculation in MVBs is induced by the EGF-R and is mediated by phosphorylated annexin I.
Futter
C E
CE
MRC Laboratory for Molecular Cell Biology, University College, London.
Felder
S
S
Schlessinger
J
J
Ullrich
A
A
Hopkins
C R
CR
D016475:3T3 Cells; D000818:Animals; D017305:Annexin A1 / Q000378:metabolism*; D001692:Biological Transport; D002118:Calcium / Q000378:metabolism; D002451:Cell Compartmentation; D002458:Cell Fractionation; D002462:Cell Membrane / Q000378:metabolism; D004705:Endocytosis; D011992:Endosomes / Q000378:metabolism*; D066246:ErbB Receptors / Q000378:metabolism*; D006801:Humans; D051379:Mice; D015388:Organelles / Q000378:metabolism*; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism
D017305:Annexin A1; D011994:Recombinant Proteins; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases; D002118:Calcium