EEA1, AN EARLY ENDOSOME-ASSOCIATED PROTEIN - EEA1 IS A CONSERVED ALPHA-HELICAL PERIPHERAL MEMBRANE-PROTEIN FLANKED BY CYSTEINE FINGERS AND CONTAINS A CALMODULIN-BINDING IQ MOTIF

Early endosomes are cellular compartments receiving endocytosed materi al and sorting them for vesicular transport to late endosomes and lyso somes or for recycling to the plasma membrane. We have cloned a human cDNA encoding an evolutionarily conserved 180-kDa protein on early end osomes named EEA1 (Early Endosome Antigen1). EEA1 is associated with e arly endosomes since it co-localizes by immunofluorescence with the tr ansferrin receptor and with Rab5 but not with Rab7. Immunoelectron mic roscopy shows that it is associated with tubulovesicular early endosom es containing internalized bovine serum albumin-gold. EEA1 is a hydrop hilic peripheral membrane protein present in cytosol and membrane frac tions. It partitions in the aqueous phase after Triton X-114 solubiliz ation and is extracted from membranes by 0.3 M NaCl. It is a predomina ntly cu-helical protein sharing 17-20% sequence identity with the myos ins and contains a calmodulin-binding IQ motif. It is flanked by metal -binding, cysteine ''finger'' motifs. The COOH-terminal fingers, Cys-X (2)-Cys-X(12)-Cys-X(2)-Cys and Cys-X(2)-Cys-X(16)-Cys-X(2)-Cys, are pr esent within a region that is strikingly homologous with Saccharomyces cerevisiae FAB1 protein required for endocytosis and with Caenarhabdi tis elegans ZK632. These fingers also show limited conservation with S . cerevisiae VAC1, Vps11, and Vps18p proteins implicated in vacuolar t ransport. We propose that EEA1 is required for vesicular transport of proteins through early endosomes and that its finger motifs are requir ed for this activity.