The finding that patients and mice bearing autoantibodies directed against early endosome antigen 1 (EEA1) develop neurological signs and deficits prompted an investigation of EEA1 distribution, localization, and interaction with synaptic proteins found in neural tissues. We detected EEA1 in a variety of neural tissues and in cells of neural origin where it co-localized with SNAP-25. The interaction between EEA1 and SNAP-25 was dependent on the leucine zipper and a newly identified methyl-accepting domain of EEA1. The C-terminal zinc-binding FYVE finger motif (EEA1(1271-1411)) of EEA1 also interacted with native SNAP-25 but only in the presence of 100microM Ca(2+). In contrast, EEA1 did not bind to cysteine string protein or synapsin in these binding assays. These results suggest that EEA1 is involved in neuronal synaptic vesicle function and axonal transport and growth. EEA1 may undergo calcium-dependent conformational changes that are required for binding to SNAP-25.