Ca(2+)/calmodulin-dependent protein kinase II (CaMK-II) isozyme variability is the result of alternative usage of variable domain sequences. Isozyme expression is cell type-specific to transduce the appropriate Ca(2+) signals. We have determined the subcellular targeting domain of delta(E) CaMK-II, an isozyme that induces neurite outgrowth, and of a structurally similar isozyme, gamma(C) CaMK-II, which does not induce neurite outgrowth. delta(E) CaMK-II co-localizes with filamentous actin in the perinuclear region and in cellular extensions. In contrast, gamma(C) CaMK-II is uniformly cytosolic. Constitutively active delta(E) CaMK-II induces F-actin-rich extensions, thereby supporting a functional role for its localization. C-terminal constructs, which lack central variable domain sequences, can oligomerize and localize like full-length delta(E) and gamma(C) CaMK-II. Central variable domains themselves are monomeric and have no targeting capability. The C-terminal 95 residues of delta CaMK-II also has no targeting capability but can efficiently oligomerize. These findings define a targeting domain for gamma and delta CaMK-IIs that is in between the central variable and association domains. This domain is responsible for the subcellular targeting differences between gamma and delta CaMK-IIs.