Flexible Players within the Sheaths: The Intrinsically Disordered Proteins of Myelin in Health and Disease.
ABSTRACT: Myelin ensheathes selected axonal segments within the nervous system, resulting primarily in nerve impulse acceleration, as well as mechanical and trophic support for neurons. In the central and peripheral nervous systems, various proteins that contribute to the formation and stability of myelin are present, which also harbor pathophysiological roles in myelin disease. Many myelin proteins have common attributes, including small size, hydrophobic segments, multifunctionality, longevity, and regions of intrinsic disorder. With recent advances in protein biophysical characterization and bioinformatics, it has become evident that intrinsically disordered proteins (IDPs) are abundant in myelin, and their flexible nature enables multifunctionality. Here, we review known myelin IDPs, their conservation, molecular characteristics and functions, and their disease relevance, along with open questions and speculations. We place emphasis on classifying the molecular details of IDPs in myelin, and we correlate these with their various functions, including susceptibility to post-translational modifications, function in protein-protein and protein-membrane interactions, as well as their role as extended entropic chains. We discuss how myelin pathology can relate to IDPs and which molecular factors are potentially involved.
Project description:All vertebrate nervous systems, except those of agnathans, make extensive use of the myelinated fiber, a structure formed by coordinated interplay between neuronal axons and glial cells. Myelinated fibers, by enhancing the speed and efficiency of nerve cell communication allowed gnathostomes to evolve extensively, forming a broad range of diverse lifestyles in most habitable environments. The axon-covering myelin sheaths are structurally and biochemically novel as they contain high portions of lipid and a few prominent low molecular weight proteins often considered unique to myelin. Here we searched genome and EST databases to identify orthologs and paralogs of the following myelin-related proteins: (1) myelin basic protein (MBP), (2) myelin protein zero (MPZ, formerly P0), (3) proteolipid protein (PLP1, formerly PLP), (4) peripheral myelin protein-2 (PMP2, formerly P2), (5) peripheral myelin protein-22 (PMP22) and (6) stathmin-1 (STMN1). Although widely distributed in gnathostome/vertebrate genomes, neither MBP nor MPZ are present in any of nine invertebrate genomes examined. PLP1, which replaced MPZ in tetrapod CNS myelin sheaths, includes a novel 'tetrapod-specific' exon (see also Möbius et al., 2009). Like PLP1, PMP2 first appears in tetrapods and like PLP1 its origins can be traced to invertebrate paralogs. PMP22, with origins in agnathans, and STMN1 with origins in protostomes, existed well before the evolution of gnathostomes. The coordinated appearance of MBP and MPZ with myelin sheaths and of PLP1 with tetrapod CNS myelin suggests interdependence - new proteins giving rise to novel vertebrate structures.
Project description:Myelin is a multilayered membrane that ensheathes axonal fibers in the vertebrate nervous system, allowing fast propagation of nerve action potentials. It contains densely packed lipids, lacks an actin-based cytocortex, and requires myelin basic protein (MBP) as its major structural component. This protein is the basic constituent of the proteinaceous meshwork that is localized between adjacent cytoplasmic membranes of the myelin sheath. Yet, it is not clear how MBP influences the organization and dynamics of the lipid constituents of myelin. Here, we used optical stimulated emission depletion super-resolution microscopy in combination with fluorescence correlation spectroscopy to assess the characteristics of diffusion of different fluorescent lipid analogs in myelin membrane sheets of cultured oligodendrocytes and in micrometer-sized domains that were induced by MBP in live epithelial PtK2 cells. Lipid diffusion was significantly faster and less anomalous both in oligodendrocytes and inside the MBP-rich domains of PtK2 cells compared with undisturbed live PtK2 cells. Our data show that MBP reorganizes lipid diffusion, possibly by preventing the buildup of an actin-based cytocortex and by preventing most membrane proteins from entering the myelin sheath region. Yet, in contrast to myelin sheets in oligodendrocytes, the MBP-induced domains in epithelial PtK2 cells demonstrate no change in lipid order, indicating that segregation of long-chain lipids into myelin sheets is a process specific to oligodendrocytes.
Project description:Myelin ensheathes axons to allow rapid propagation of action potentials and proper nervous system function. In the peripheral nervous system, Schwann cells (SCs) radially sort axons into a 1:1 relationship before wrapping an axonal segment to form myelin. SC myelination requires the adhesion G protein-coupled receptor GPR126, which undergoes autoproteolytic cleavage into an N-terminal fragment (NTF) and a seven-transmembrane-containing C-terminal fragment (CTF). Here we show that GPR126 has domain-specific functions in SC development whereby the NTF is necessary and sufficient for axon sorting, whereas the CTF promotes wrapping through cAMP elevation. These biphasic roles of GPR126 are governed by interactions with Laminin-211, which we define as a novel ligand for GPR126 that modulates receptor signaling via a tethered agonist. Our work suggests a model in which Laminin-211 mediates GPR126-induced cAMP levels to control early and late stages of SC development.
Project description:The 18.5 kDa isoform of myelin basic protein (MBP) is the predominant form in adult human central nervous system myelin. It is an intrinsically disordered protein that functions both in membrane adhesion, and as a linker connecting the oligodendrocyte membrane to the underlying cytoskeleton; its specific interactions with calmodulin and SH3-domain containing proteins suggest further multifunctionality in signaling. Here, we have used multidimensional heteronuclear nuclear magnetic resonance spectroscopy to study the conformational dependence on environment of the protein in aqueous solution (100 mM KCl) and in a membrane-mimetic solvent (30% TFE-d(2)), particularly to analyze its secondary structure using chemical shift indexing, and to investigate its backbone dynamics using (15)N spin relaxation measurements. Collectively, the data revealed three major segments of the protein with a propensity toward alpha-helicity that was stabilized by membrane-mimetic conditions: T33-D46, V83-T92, and T142-L154 (murine 18.5 kDa sequence numbering). All of these regions corresponded with bioinformatics predictions of ordered secondary structure. The V83-T92 region comprises a primary immunodominant epitope that had previously been shown by site-directed spin labeling and electron paramagnetic resonance spectroscopy to be alpha-helical in membrane-reconstituted systems. The T142-L154 segment overlapped with a predicted calmodulin-binding site. Chemical shift perturbation experiments using labeled MBP and unlabeled calmodulin demonstrated a dramatic conformational change in MBP upon association of the two proteins, and were consistent with the C-terminal segment of MBP being the primary binding site for calmodulin.
Project description:During development, oligodendrocytes contact and wrap neuronal axons with myelin. Similarly to neurons and synapses, excess myelin sheaths are produced and selectively eliminated, but how elimination occurs is unknown. Microglia, the resident immune cells of the central nervous system, engulf surplus neurons and synapses. To determine whether microglia also prune myelin sheaths, we used zebrafish to visualize and manipulate interactions between microglia, oligodendrocytes, and neurons during development. We found that microglia closely associate with oligodendrocytes and specifically phagocytose myelin sheaths. By using a combination of optical, genetic, chemogenetic, and behavioral approaches, we reveal that neuronal activity bidirectionally balances microglial association with neuronal cell bodies and myelin phagocytosis in the optic tectum. Furthermore, multiple strategies to deplete microglia resulted in oligodendrocytes maintaining excessive and ectopic myelin. Our work reveals a neuronal activity-regulated role for microglia in modifying developmental myelin targeting by oligodendrocytes.
Project description:BACKGROUND: During the development of the central nervous system, oligodendrocytes generate large amounts of myelin, a multilayered insulating membrane that ensheathes axons, thereby allowing the fast conduction of the action potential and maintaining axonal integrity. Differentiation of oligodendrocytes to myelin-forming cells requires the downregulation of RhoA GTPase activity. RESULTS: To gain insights into the molecular mechanisms of oligodendrocyte differentiation, we performed microarray expression profiling of the oligodendroglial cell line, Oli-neu, treated with the Rho kinase (ROCK) inhibitor, Y-27632 or with conditioned neuronal medium. This resulted in the identification of the transmembrane protein 10 (Tmem10/Opalin), a novel type I transmembrane protein enriched in differentiating oligodendrocytes. In primary cultures, Tmem10 was abundantly expressed in O4-positive oligodendrocytes, but not in oligodendroglial precursor cells, astrocytes, microglia or neurons. In mature oligodendrocytes Tmem10 was enriched in the rims and processes of the cells and was only found to a lesser extent in the membrane sheets. CONCLUSION: Together, our results demonstrate that Tmem10 is a novel marker for in vitro generated oligodendrocytes.
Project description:Myelin is best known for its role in increasing the conduction velocity and metabolic efficiency of long-range excitatory axons. Accordingly, the myelin observed in neocortical gray matter is thought to mostly ensheath excitatory axons connecting to subcortical regions and distant cortical areas. Using independent analyses of light and electron microscopy data from mouse neocortex, we show that a surprisingly large fraction of cortical myelin (half the myelin in layer 2/3 and a quarter in layer 4) ensheathes axons of inhibitory neurons, specifically of parvalbumin-positive basket cells. This myelin differs significantly from that of excitatory axons in distribution and protein composition. Myelin on inhibitory axons is unlikely to meaningfully hasten the arrival of spikes at their pre-synaptic terminals, due to the patchy distribution and short path-lengths observed. Our results thus highlight the need for exploring alternative roles for myelin in neocortical circuits.
Project description:The number of myelin sheaths made by individual oligodendrocytes regulates the extent of myelination, which profoundly affects central nervous system function. It remains unknown when, during their life, individual oligodendrocytes can regulate myelin sheath number in vivo. We show, using live imaging in zebrafish, that oligodendrocytes make new myelin sheaths during a period of just 5 hr, with regulation of sheath number after this time limited to occasional retractions. We also show that activation and reduction of Fyn kinase in oligodendrocytes increases and decreases sheath number per cell, respectively. Interestingly, these oligodendrocytes also generate their new myelin sheaths within the same period, despite having vastly different extents of myelination. Our data demonstrate a restricted time window relative to the lifetime of the individual oligodendrocyte, during which myelin sheath formation occurs and the number of sheaths is determined.
Project description:Intrinsically disordered proteins (IDPs) or unstructured segments within proteins play an important role in cellular physiology and pathology. Low cellular concentration, multiple binding partners, frequent post-translational modifications and the presence of multiple conformations make it difficult to characterize IDP interactions in intact cells. We used peptide aptamers selected by using the yeast-two-hybrid scheme and in-cell NMR to identify high affinity binders to transiently structured IDP and unstructured segments at atomic resolution. Since both the selection and characterization of peptide aptamers take place inside the cell, only physiologically relevant conformations of IDPs are targeted. The method is validated by using peptide aptamers selected against the prokaryotic ubiquitin-like protein, Pup, of the mycobacterium proteasome. The selected aptamers bind to distinct sites on Pup and have vastly different effects on rescuing mycobacterial proteasome substrate and on the survival of the Bacille-Calmette-Guèrin, BCG, strain of M. bovis. This technology can be applied to study the elusive action of IDPs under near physiological conditions.
Project description:BACKGROUND:Intrinsically disordered proteins or regions (IDPs or IDRs) lack stable structures in solution, yet often fold upon binding with partners. IDPs or IDRs are highly abundant in all proteomes and represent a significant modification of sequence ? structure ? function paradigm. The Protein Data Bank (PDB) includes complexes containing disordered segments bound to globular proteins, but the molecular mechanisms of such binding interactions remain largely unknown. RESULTS:In this study, we present the results of various disorder predictions on a nonredundant set of PDB complexes. In contrast to their structural appearances, many PDB proteins were predicted to be disordered when separated from their binding partners. These predicted-to-be-disordered proteins were observed to form structures depending upon various factors, including heterogroup binding, protein/DNA/RNA binding, disulfide bonds, and ion binding. CONCLUSIONS:This study collects many examples of disorder-to-order transition in IDP complex formation, thus revealing the unusual structure-function relationships of IDPs and providing an additional support for the newly proposed paradigm of the sequence ? IDP/IDR ensemble ? function.