Tension Sensor Based on Fluorescence Resonance Energy Transfer Reveals Fiber Diameter-Dependent Mechanical Factors During Myelination.

Oligodendrocytes (OLs) form a myelin sheath around neuronal axons to increase conduction velocity of action potential. Although both large and small diameter axons are intermingled in the central nervous system (CNS), the number of myelin wrapping is related to the axon diameter, such that the ratio of the diameter of the axon to that of the entire myelinated-axon unit is optimal for each axon, which is required for exerting higher brain functions. This indicates there are unknown axon diameter-dependent factors that control myelination. We tried to investigate physical factors to clarify the mechanisms underlying axon diameter-dependent myelination. To visualize OL-generating forces during myelination, a tension sensor based on fluorescence resonance energy transfer (FRET) was used. Polystyrene nanofibers with varying diameters similar to neuronal axons were prepared to investigate biophysical factors regulating the OL-axon interactions. We found that higher tension was generated at OL processes contacting larger diameter fibers compared with smaller diameter fibers. Additionally, OLs formed longer focal adhesions (FAs) on larger diameter axons and shorter FAs on smaller diameter axons. These results suggest that OLs respond to the fiber diameter and activate mechanotransduction initiated at FAs, which controls their cytoskeletal organization and myelin formation. This study leads to the novel and interesting idea that physical factors are involved in myelin formation in response to axon diameter.

Social Defeat Stress in Adolescent Mice Induces Depressive-like Behaviors with Reduced Oligodendrogenesis.

Strong stress related to adverse experiences during adolescence can cause mental disorders, as well as affecting brain structure and function. However, the underlying neurobiological mechanisms remain largely unknown. To investigate whether stress induced by adverse experience during adolescence affects oligodendrocyte (OL) remodeling, social defeat stress was applied to 6-week-old adolescent mice for 10 days, followed by behavioral tests and assessments of oligodendrogenesis. Socially defeated mice showed depressive-like behaviors in behavioral experiments. Stress led to a decrease in the number of newly born OLs in the anterior cortical region and the number of proteolipid protein-positive mature OLs in the corpus callosum and posterior cerebral cortex. Fewer bromodeoxyuridine-incorporated CC1-positive mature OLs were observed in these regions in socially defeated mice. To assess whether decreased oligodendrogenesis caused by social defeat stress is related to depressive-like symptoms under stress, clemastine, a drug that induces OL generation, was administered to socially defeated adolescent mice, resulting in the rescue of the behavioral abnormalities accompanied by increased oligodendrogenesis. These findings suggest that oligodendrogenesis in adverse environments during adolescence plays a role in psychiatric disorders, and clemastine may provide a potential therapeutic drug for adolescent mental disorders, targeting OLs.

N,N-Dimethylaminopyrene as a fluorescent affinity mass tag for ligand-binding mode analysis.

Elucidation of the binding mode of protein-ligand interactions provides insights for the design of new pharmacological tools and drug leads. Specific labeling of target proteins with chemical probes, in which the ligands are conjugated with reacting and detecting groups, can establish the binding positions of ligands. Label-assisted laser desorption/ionization mass spectrometry (LA-LDI MS) is a promising detection method to selectively detect labeled molecules. However, previous LDI MS tags, such as nitrogen-substituted pyrenes, had problems with low sensitivity and stability. Here we show 6-N,N-dimethylaminopyrene (dmpy) as a versatile mass tag, which was detected at an amount of 0.1 fmol by LA-LDI MS and applicable for MS/MS analysis. By using ligand-dissociation-type dmpy probes and affinity purification with a polystyrene gel, we demonstrated that dmpy-labeled peptides were predominantly detected by MALDI MS. Our dmpy-probe-labeling method might be highly useful for determining the target biomacromolecules of various ligands and their binding sites.

Specific protein-labeling and ligand-binding position analysis with amidopyrene probes as LDI MS tags.

To readily analyze the binding mode of protein-ligand interactions, we developed ligand-bound-type and ligand-dissociation-type probes having 6-amidopyrene (apy) as a detecting group. Matrix- and label-assisted laser desorption/ionization mass spectrometry (MALDI and LA-LDI MS) analyses and a covalent docking simulation using these probes precisely determined the binding position of the ligand biotin on the target protein avidin (RMSD = 0.786 and 0.332 Å). Our apy-probe-labeling method may be useful for determining the unknown ligand-binding sites of various target proteins.