Myelin is a multilamellar, lipid-rich membrane that insulates axons in the vertebrate nervous system to facilitate the rapid conduction of action potentials. Myelin is generated by specialized glial cells - Schwann cells (SCs) in the peripheral nervous system and oligodendrocytes (OLs) in the central nervous systems. Both cell types form myelin by extending and iteratively wrapping their plasma membranes around axons, and glial cell cytoplasm is extruded to ultimately form the myelin sheath. In addition, myelinating gli promote neuronal health by providing vital trophic support to the axons they ensheath. Impaired myelination is the cause of many severe neurological disorders such as multiple sclerosis and Charcot-Marie-Tooth disease, and loss of myelin can lead to neuronal loss and eventual paralysis. Although myelin is essential for human life, the molecular mechanisms that underlie glial cell development and myelination are poorly understood. To this end, we seek to identify and characterize novel regulators of myelination. We recently determined that the adhesion-GPCR (aGPCR) Gpr126 is essential for SC myelination. Adhesion-GPCRs are characterized by the presence of a large N-terminus often enriched for domains associated with cell-cell/cell-matrix interactions. The dual roles of aGPCRs in facilitating cell-cell interactions and their intracellular signaling capacity, in addition to the importance of Gpr126 for SC myelination, led us to hypothesize that additional aGPCRs may regulate glial cell development and myelination. We have determined that Gpr56, an aGPCR related to Gpr126, is highly expressed in myelinating glia, and our preliminary analyses of zebrafish and mouse mutants indicate that Gpr56 is an important regulator of OL and SC myelination. In this proposal, we seek to define the role of Gpr56 in myelinating glia. In the first aim, I will test the hypothesis that Gpr56 is required for oligodendrocyte development and myelination using two gpr56 zebrafish mutants I generated using Transcription Activator-Like Effector Nucleases. In addition to traditional molecular and ultrastructural techniques, I will take advantage of the optical clarity of zebrafish by performing time-lapse imaging to determine how loss of gpr56 function affects glial cell behavior in living embryos. In the second aim, I will test the hypothesis that Gpr56 is required fo SC development and myelin maintenance. To this end, we will examine the consequences of Gpr56 loss of function using immunohistochemical and ultrastructural approaches in both zebrafish and mouse mutants. In the third aim, we will employ a structure-function based strategy to determine which functional domains (signaling versus adhesion) are necessary for Gpr56 regulation of OL and SC myelination. Specifically, we will test the ability of single functional domains (N- terminus versus C-terminus) to rescue gpr56 mutant phenotypes. Together, these experiments will define the role of Gpr56 in SC and OL myelination and myelin maintenance and will broaden our basic understanding of aGPCR biology and the molecular mechanisms that control myelination.
Genetic defects that cause aberrant myelin development or demyelination are incompatible with a high quality of human life, but therapies that prevent demyelination and/or promote robust remyelination are either inadequate or nonexistent. Here, we propose to determine how the G protein-coupled receptor Gpr56 regulates myelin formation in development and myelin maintenance throughout adulthood. These studies will dissect the genetic and molecular mechanisms that control myelination, which is essential for developing new strategies for preventing devastating demyelinating diseases.
