Precise navigation of axons to their targets is critical for establishing proper neuronal networks during development. Axon elongation, whereby axons extend far beyond the site of initiation to reach their target cells, is an essential step in this process, but the precise molecular pathways that regulate axon growth remain uncharacterized. Here we show that 14‐3‐3/14‐3‐3ς proteins—adaptor proteins that modulate diverse cellular processes including cytoskeletal dynamics—play a critical role in Xenopus retinal ganglion cell (RGC) axon elongation in vivo and in vitro. We have identified the expression of 14‐3‐3/14‐3‐3ς transcripts and proteins in retinal growth cones, with higher levels of expression occurring during the phase of rapid pathway extension. Competitive inhibition of 14‐3‐3/14‐3‐3ς by expression of a genetically encoded peptide, R18, in RGCs resulted in a marked decrease in the length of the initial retinotectal projection in vivo and a corresponding decrease in axon elongation rate in vitro (30–40%). Furthermore, 14‐3‐3/14‐3‐3ς (R1) co‐localized with Xenopus actin depolymerizing factor (ADF)/cofilin (XAC) in RGC growth cones. Inhibition of 14‐3‐3/14‐3‐3ς function with either R18 or morpholinos reduced the level of inactive pXAC and increased the sensitivity to collapse by the repulsive cue, Slit2. Collectively, these results demonstrate that14‐3‐3/14‐3‐3ς participates in the regulation of retinal axon elongation, in part by modulating XAC activity. © 2011 Wiley Periodicals, Inc. Develop Neurobiol 72: 600–614, 2012