ERAD-derived preprotein transport across the second outermost plastid membrane of diatoms.

The diatom Phaeodactylum tricornutum harbors a plastid that is surrounded by four membranes and evolved by way of secondary endosymbiosis. Like land plants, most of its plastid proteins are encoded as preproteins on the nuclear genome of the host cell and are resultantly redirected into the organelle. Because two more membranes are present in diatoms than the one pair surrounding primary plastids, the targeting situation is obviously different and more complex. In this work, we focus on preprotein transport across the second outermost plastid membrane -- an issue that was experimentally inaccessible until now. We provide first indications that our hypothesis of an ERAD (ER-associated degradation)-derived preprotein transport system might be correct. Our data demonstrate that the symbiont-specific Der1 proteins, sDer1-1 and sDer1-2, form an oligomeric complex within the second outermost membrane of the complex plastid. Moreover, we present first evidence that the complex interacts with transit peptides of preproteins being transported across this membrane into the periplastidal compartment but not with transit peptides of stromal-targeted proteins. Thus, the sDer1 complex might have an additional role in discriminating preproteins that are transported across the two outermost membranes from preproteins directed across all four membranes of the complex plastid. Altogether, our studies of the symbiont-specific ERAD-like machinery of diatoms suggest that a preexisting cellular machinery was recycled to fulfill a novel function during the transition of a former free-living eukaryote into a secondary endosymbiont.