PLASTOCYANIN AND THE 33-KDA SUBUNIT OF THE OXYGEN-EVOLVING COMPLEX ARE TRANSPORTED INTO THYLAKOIDS WITH SIMILAR REQUIREMENTS AS PREDICTED FROM PATHWAY SPECIFICITY

Plastocyanin and the 33-kDa subunit of the oxygen-evolving complex (OE 33) are two of several thylakoid lumen-located proteins that are made in the cytosol, imported into chloroplasts, and subsequently transport ed into thylakoids. Recently, competition studies showed that there ar e two pathways for protein transport into the thylakoid lumen and that plastocyanin and OE33 are on the same pathway (Cline, K., Henry, R., Li, C., and Yuan, J. (1993) EMBO J. 12, 4105-4114). Our expectation is that transport requirements reflect the steps of the process and that proteins on the same pathway share similar requirements. Unfortunatel y, the transport requirements for plastocyanin and OE33 are not well e stablished. Here, we investigated transport in a reconstituted system with isolated thylakoids. Efficient transport of OE33 and plastocyanin was only obtained when stromal extract was included in the assay. Hea t or protease treatment of stromal extract eliminated its ability to s timulate transport. Transport was abolished by treatments designed to deplete ATP or to prevent its formation and was greatly reduced in the presence of ionophores that dissipate the trans thylakoidal proton gr adient. These results show that transport of OE33 and plastocyanin req uires ATP and is stimulated by stromal protein(s) and the trans-thylak oidal proton gradient. Taken together, these and previous results sugg est that there are two mechanistically distinct pathways for protein t ransport into the thylakoid lumen.