The endosymbiotic organelles, mitochondria and chloroplasts in eukaryotic cells fulfil essential biochemical duties and at the same time represent a prominent proportion of the cellular proteom. The biogenesis requires the coordinated expression of organellar and nuclear genomes and a coordinated synthesis, transport and assembly of proteins in time, space and developmental specific manner. We have learned in recent years that a multitude of factors either cytosolic or localized in the target organelles are involved in the process of high fidelity subcellular distribution, targeting and translocation. Surprisingly, each compartment in a eukaryotic cell has developed multiple translocation and routing pathways each employing again numerous subunits, many of which do not function as a single entity, but only as a part of complex sophisticated molecular machines. Though in many cases we know the parts, which assemble the machine, we do not know how they are put together to form the machine and how each part functions in the context of the overall motor. Surprisingly the endosymbiotic organelles mitochondria and chloroplasts have developed quite distinct strategies to target and translocate proteins. This could have been necessary, because chloroplasts entered the cell at a later point in evolution than mitochondria. We plan to address these questions using also the single molecule spectroscopy tools provided by research area A.