molecular analysis of rapid endocytosis and plasma membrane recycling in the model eukaryote Trypanosoma brucei

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African trypanosomes are extracellular parasites that cause human sleeping sickness. Key to their success has been the ability to endocytose essential macromolecules from the mammalian host without exposing the machinery involved to the attention of the immune system, which has led to the evolution of an extraordinary system for membrane traffic. Uniquely, endocytosis in trypanosomes is highly polarized and occurs solely at the terminus of the exocytotic arm of membrane traffic: the flagellar pocket. Although this site represents less than 5% of the total surface, membrane traffic here far exceeds that of any other eukaryote. Concurrent with this huge traffic the pathway allows the differential sorting, internalization and recycling of abundant glycosylphosphoinositol anchored proteins. The molecular basis for this only recently described pathway remains elusive. We propose a systematic approach to the molecular characterization of the T. brucei endosome that exploits RNA interference, high-resolution 3D microscopy, novel methodology for isolation of endosomes, isotope-tagging methods and mass spectrometry. This powerful screen focuses on important, general questions regarding endocytosis and recycling of lipid-anchored proteins. The molecular quantification of the trypanosome endocytosis machinery not only presents a unique opportunity to deepen our understanding of membrane traffic at a fundamental level, but may also challenge a fatal pathogen.