The vacuolar system of plants is a key element of plant growth and development and fulfills many other functions. Plant cells can have two different vacuoles, requiring different sorting systems, with different signals, receptors, transport vesicles, etc. The vacuolar systems of higher plants have evolved from simpler ancestors, which might have been preserved in lower plants. This evolution is reflected in the gene families involved in vacuole biogenesis. Analysis of knock-out or silencing mutants and their complementation will allow to understand the and the capacity of higher plants to adapt to their environment. We intend to clarify the role of members of two receptor families in targeting to different vacuoles and we will focus mainly on the RMR family, the role of which is currently disputed. Some results point to a direct role as receptor, while other results suggest a more basic role in all pathways to the vacuoles. We will develop a new mutant screen for genes implicated in vacuolar trafficking, with an emphasis on the root tissues of higher plants. We will continue to develop the moss Physcomitrella patens as a model system for studies of the secretory pathway in plants, taking advantages of the exceptional genetic tools available, in particular the possibilities of targeted gene knock-outs (even multiple), of gene replacement and targeted mutagenesis. We will thus be able to compare the two very different types of land plants, to determine which elements of the plant specific aspects of the secretory pathway date back to the conquest of land by plants and which may have developed because of the increasing tissue specialization, particularly in flowering plants.