ENDOSOMAL LOCALIZATION OF THE AUTOANTIGEN EEA1 IS MEDIATED BY A ZINC-BINDING FYVE FINGER

EEA1, a 162-kDa autoantigen associated with subacute cutaneous systemi c lupus erythematosus, is a coiled-coil protein localized to early end osomes and cytosol. At its C terminus, the protein contains a cysteine rich motif, which is shared with Vps27, Fab1, and Vac1, yeast protein s implicated in membrane traffic (Mu, F. T., Callaghan, J. M., Steele- Mortimer, O., Stenmark, H., Parton, R. G., Campbell, P. L., McCluskey, J., Yeo, J. P., Tock, E. P., and Toh, B. H. (1995) J. Biol. Chem. 270 , 13503-13511). Here we show that this motif constitutes a genuine zin c binding domain, which we term the FYVE finger (based on the first le tters of four proteins containing this motif). Profile based data base searches identified the FYVE finger in 11 distinct proteins. The FYVE finger containing C terminus of EEA1 was found to bind 2 mol equivale nts of Zn2+. Mutations of conserved histidine and cysteine residues in the FYVE motif independently reduced zinc binding to 1 mol equivalent . Confocal immunofluorescence microscopy of transfected HEp2 cells rev ealed that the C-terminal part (residues 1277-1411) of EEA1 colocalize s extensively with a GTPase deficient mutant of the early endosomal GT Pase Rab5, while deletion of the FYVE finger or mutations that interfe re with zinc binding cause a cytosolic localization. These results imp licate the FYVE finger in the specific localization of EEA1 to endosom es.