We report here a novel intracellular localization and function of Tau proteins in cultured cerebellar neurons. Immunofluorescence staining of detergent-extracted cytoskeletons with antibodies specific for Tau proteins revealed intense labeling of growth cone microtubules. Besides, suppression of Tau by antisense oligonucleotide treatment results in the complete disappearance of antigen 13H9, a specific growth cone component with properties of microfilament- and microtubule-associated protein [Goslin et al., 1989: J. Cell Biol. 109:1621-1631], from its normal intracellular location. This phenomenon is unique to neurite-bearing cells, is not associated with the disappearance of microtubules from growth cones, and is not reversed by taxol, a microtubule-stabilizing agent. In addition, Tau-suppressed neurons display a significant reduction in growth cone area and fillopodial number; on the contrary, fillopodial length increases significantly. The alterations in growth cone morphology are accompanied by considerable changes in the phalloidin staining of assembled actin. Taken together, the present results suggest that in developing neurons Tau proteins participate in mediating interactions between elements of the growth cone cytoskeleton important for maintaining the normal structural organization of this neuritic domain.