Throughout the last thirty years, research on Rab GTPase proteins has brought light to their pivotal role in the regulation of several steps of intracellular trafficking. However, recent findings indicate that Rab proteins not only regulate individual transport steps, but they also function in a coordinated fashion in Rab GTPase cascades. Such cascades appear to be crucial for orchestrating sequential steps along transport pathways and for ensuring continuity in membrane traffic. Rab cascades have been found in the endocytic pathway of mammalian cells (Rab5/Rab7) as well as in the secretory pathway of yeast (Ypt32p/Sec4p) but none have been, so far, identified in the secretory pathways of the former. Therefore, the main goal of this research project is to identify Rab GTPase cascades in the post-Golgi trafficking of mammalian cells. By using a multidisciplinary approach, we propose to: 1) establish a comprehensive list of Rab GTPases involved in the post-Golgi trafficking from the trans-Golgi network to the plasma membrane and to the endolysosomal compartments; 2) determine the temporal recruitment of Rab GTPases to the post-Golgi vesicles associated with each one of the pathways and identify Rab cascades; 3) identify the Guanine nucleotide Exchange Factor (GEF) proteins involved in the cascades previously found. Achieving this work plan will increase our knowledge on the regulation of sequential trafficking steps, essential for the full understanding of human pathologies, like cancer and neurological diseases. Therefore, we will be contributing to the scientific European excellence by reaching one of the main goals of the Seventh Framework Programme regarding Health.