The myelin sheath is a highly specialized membranous structure that surrounds axons of the central and peripheral nervous systems and is essential for the normal saltatory conduction of the electrical impulse along the axon. The cells responsible for the formation of myelin in the brain are called oligodendrocytes. Some of the myelin proteins are synthesized in the body of the oligodendrocyte and subsequently transported by small vesicles to the place where the myelin is formed. Similarly, myelin components can be transported from the myelin sheath to other parts of the oligodendrocytes by means of small vesicles. The formation, maintenance and remodeling of the myelin sheath depend at least in part on the intracellular transport of vesicles. The transport of vesicles is regulated by controlling both vesicle formation and fusion. At present little is known about the mechanisms in oligodendrocytes that regulate this traffic, because of the previous absence of definitive experimental approaches to sort out the various signalling components that function in living oligodendrocytes. We were able to design an approach that allows the visualization of different pathways of vesicle transport in living oligodendrocytes. We isolated from oligodendrocytes a novel gene that codifies for a protein, designated RabX, which is a member of a family of proteins that are known to regulated specific vesicle trafficking pathways. The Rab family of proteins regulates intracellular transport of vesicles controlling both the formation of vesicles in the membrane donor compartment and the fusion/targeting of vesicles in the acceptor compartment. Additionally, Rab proteins appear to be involved in the movement of the vesicles from the donor to the acceptor compartment, which is facilitated by elements of the cell cytoskeleton. The functions of Rab proteins to regulate different steps in intracellular vesicle transport depend on their ability to alternate between two conformational states, inactive (GDP-bound) and active (GTP-bound). The capacity to change conformational state relies on GTPase activity. Our research will be focus in the study of RabX GTPase activity and in the analysis of the movement in living cells of vesicles containing RabX. Using molecular biological techniques RabX will be attached to EYFP (a fluorescent protein). This chimera protein (RabX-EYFP) will be express in different cells including oligodendrocytes, and since it is fluorescent we will able to see the intracellular compartment that contain Rab0-EYFP by fluorescence microscopy. By taking pictures of the cells at different times we will able to study the movement of vesicles that contain RabX-EYFP. It is expected that this research will result in a better understanding of how RabX regulates the intracellular transport of vesicles in the oligodendrocytes. This information is essential for knowing how myelin formation and remodeling take place. We believe that this basic knowledge will help in the design of novel treatments for recovering the myelin lost during demyelinating disorders, such as multiple sclerosis and spinal cord injury.
