The malarial "apicoplast" derived from an algal plastid, has stimulated interest for its novel evolutionary biology and potential as a drug target. An endoplasmic reticulum-type signal sequence followed by a plastid targeting sequence are required to target proteins to the apicoplast but the pathway by which proteins are transported to the organelle is unknown. By stage regulating the expression of transgenes we show that early (0-12 h) in the parasite's development in red cells, newly synthesized green fluorescent protein that contains the plastid targeting sequence (plastid targeting sequence-green fluorescent protein (PTS-GFP)) is recruited into the parasite's secretory pathway. PTS-GFP in 0-12-h parasites is found released into the parasitophorous vacuole (PV) and in apposition with the Golgi. However, import into the apicoplast and processing to GFP does not occur until 18-36 h in development. In intermediate, 18-h parasites PTS-GFP resides in the PV. Quantitative exit of PTS-GFP from the PV and its conversion to GFP is seen at 36 h. The data suggest that: (i) import into the apicoplast is stage regulated and (ii) the PTS can signal endomembrane targeting from the PV to the apicoplast.