Transport and insertion mechanisms of plastid-encoded thylakoid membrane proteins

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The biogenesis of photosynthetic protein complexes of the chloroplast thylakoid membrane requires highly specific protein sorting, integration and assembly mechanisms of nucleus as well as plastid encoded subunits. Central steps in the biogenesis of photosystem II (PS II) are the cotranslational insertion of the plastid encoded D1 protein into the thylakoid membrane and its subsequent assembly into functional PS II. We recently established a technique to partially reconstitute the cotranslational insertion of D1 using a homologous in vitro translation system derived from pea chloroplasts. Immunological analyses of purified thylakoid membrane associated ribosomenascent chain complexes and in vitro insertion experiments revealed a role of the cpSec/Alb3 translocase and Vipp1 in D1 biogenesis. Furthermore, we recently showed that the cpftsy Arabidopsis thaliana knock-out mutant exhibits a severe defect in D1 insertion which is caused by an impaired binding of translating ribosomes to the thylakoid membrane. The aims of this proposal are (I) to identify novel components involved in cotranslational protein insertion in thylakoid membranes, (II) to dissect the protein contacts of the nascent D1 chain during translation and insertion and (III) to get insight into the mechanisms underlying targeting and attachment of ribosomenascent chain complexes to the thylakoid membrane.