Protein targeting into complex plastids : support for the trans-locator model

Plastids of diatoms are surrounded by four membranes. The outermost membrane is continuous with the endoplasmic reticulum and therefore is termed chloroplast ER (CER) membrane. The complex ultra structure of diatom plastids naturally requires more transport steps to import nucleus encoded proteins into the plastid compared to higher plant plastids which possess only two envelope membranes.Several hypothetic models for the import of preproteins into the complex plastids of diatoms are discussed. Common to all these models is the postulation of a first cotranslational transport step into the chloroplast endoplasmic reticulum lumen via the Sec61 translocon. Furthermore, all models postulate transport via a translocator in the innermost membrane similar to the Tic complex (translocon of the inner chloroplasts envelope) of higher plant plastids. The models differ, however, with respect to their explanation of transport out of the CERlumen and into the interenvelope space: either translocators, vesicles crossing the periplastidic space or putative membrane channels connecting CERlumen and the interenvelope space have been proposed. To investigate the presence of such a hypothetic connection between the CERlumen and the interenvelope space, we expressed different preproteins in the diatom Phaeodactylum tricornutum that were fused to selfassembling fragments of GFP (GFP110 and GFP11). Complementary fragments were fused to marker proteins of the CERlumen and the interenvelope space, respectively. Our data indicate that the GFP110 and GFP11 fusion proteins are located in two separate compartments which are not connected to each other.