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Light-gated ion channels and transporters have been applied to a broad array of excitable cells including neurons, cardiac myocytes, skeletal muscle cells and pancreatic β-cells in an organism to clarify their physiological and pathological roles. Nonetheless, among nonexcitable cells, only glial cells have been studied in vivo by this approach. Here, by optogenetic stimulation of a different nonexcitable cell type in the cochlea of the inner ear, we induce and control hearing loss. To our knowledge, deafness animal models using optogenetics have not yet been established. Analysis of transgenic mice expressing channelrhodopsin-2 (ChR2) induced by an oligodendrocyte-specific promoter identified this channel in nonglial cells—melanocytes—of an epithelial-like tissue in the cochlea. The membrane potential of these cells underlies a highly positive potential in a K+-rich extracellular solution, endolymph; this electrical property is essential for hearing. Illumination of the cochlea to activate ChR2 and depolarize the melanocytes significantly impaired hearing within a few minutes, accompanied by a reduction in the endolymphatic potential. After cessation of the illumination, the hearing thresholds and potential returned to baseline during several minutes. These responses were replicable multiple times. ChR2 was also expressed in cochlear glial cells surrounding the neuronal components, but slight neural activation caused by the optical stimulation was unlikely to be involved in the hearing impairment. The acute-onset, reversible and repeatable phenotype, which is inaccessible to conventional gene-targeting and pharmacological approaches, seems to at least partially resemble the symptom in a population of patients with sensorineural hearing loss. Taken together, this mouse line may not only broaden applications of optogenetics but also contribute to the progress of translational research on deafness.

Short tandem repeats are inherently unstable during DNA replication depending on repeat length, and the expansion of the repeat length in the human genome is responsible for repeat expansion disorders. Pentanucleotide AAGGG and ACAGG repeat expansions in intron 2 of the gene encoding replication factor C subunit 1 (RFC1) cause cerebellar ataxia, neuropathy, vestibular areflexia syndrome (CANVAS) and other phenotypes of late-onset cerebellar ataxia. Herein, we reveal the structural polymorphism of the RFC1 repeats associated with CANVAS in vitro. Single-stranded AAGGG repeat DNA formed a hybrid-type G-quadruplex, whereas its RNA formed a parallel-type G-quadruplex with three layers. The RNA of the ACAGG repeat formed hairpin structure comprising C-G and G-C base pairs with A:A and GA:AG mismatched repeats. Furthermore, both pathogenic repeat RNAs formed more rigid structures than those of the nonpathogenic repeat RNAs. These findings provide novel insights into the structural polymorphism of the RFC1 repeats, which may be closely related to the disease mechanism of CANVAS. [ABSTRACT FROM AUTHOR]

Light-gated ion channels and transporters have been applied to a broad array of excitable cells including neurons, cardiac myocytes, skeletal muscle cells and pancreatic β-cells in an organism to clarify their physiological and pathological roles. Nonetheless, among nonexcitable cells, only glial cells have been studied in vivo by this approach. Here, by optogenetic stimulation of a different nonexcitable cell type in the cochlea of the inner ear, we induce and control hearing loss. To our knowledge, deafness animal models using optogenetics have not yet been established. Analysis of transgenic mice expressing channelrhodopsin-2 (ChR2) induced by an oligodendrocyte-specific promoter identified this channel in nonglial cells--melanocytes--of an epithelial-like tissue in the cochlea. The membrane potential of these cells underlies a highly positive potential in a K+-rich extracellular solution, endolymph; this electrical property is essential for hearing. Illumination of the cochlea to activate ChR2 and depolarize the melanocytes significantly impaired hearing within a few minutes, accompanied by a reduction in the endolymphatic potential. After cessation of the illumination, the hearing thresholds and potential returned to baseline during several minutes. These responses were replicable multiple times. ChR2 was also expressed in cochlear glial cells surrounding the neuronal components, but slight neural activation caused by the optical stimulation was unlikely to be involved in the hearing impairment. The acuteonset, reversible and repeatable phenotype, which is inaccessible to conventional gene-targeting and pharmacological approaches, seems to at least partially resemblethe symptom in a population of patients with sensorineural hearing loss. Taken together, this mouse line may not only broaden applications of optogenetics but also contribute to the progress of translational research on deafness. [ABSTRACT FROM AUTHOR]

Tau aggregation is a defining feature of neurodegenerative tauopathies, including Alzheimer’s disease, corticobasal degeneration, and frontotemporal dementia. This aggregation involves the liquid–liquid phase separation (LLPS) of Tau, followed by its sol–gel phase transition, representing a crucial step in aggregate formation both in vitro and in vivo. However, the precise cofactors influencing Tau phase transition and aggregation under physiological conditions (e.g., ion concentration and temperature) remain unclear. In this study, we unveil that nucleic acid secondary structures, specifically RNA G-quadruplexes (rG4s), and calcium ions (Ca2+) synergistically facilitated the sol–gel phase transition of human Tau under mimic intracellular ion conditions (140 mM KCl, 15 mM NaCl, and 10 mM MgCl2) at 37 °C in vitro. In the presence of molecular crowding reagents, Tau formed stable liquid droplets through LLPS, maintaining fluidity for 24 h under physiological conditions. Notably, cell-derived RNA promoted Tau sol–gel phase transition, with rG4s emerging as a crucial factor. Surprisingly, polyanion heparin did not elicit a similar response, indicating a distinct mechanism not rooted in electrostatic interactions. Further exploration underscored the significance of Ca2+, which accumulate intracellularly during neurodegeneration, as additional cofactors in promoting Tau phase transition after 24 h. Importantly, our findings demonstrate that rG4s and Ca2+ synergistically enhance Tau phase transition within 1 h when introduced to Tau droplets. Moreover, rG4-Tau aggregates showed seeding ability in cells. In conclusion, our study illuminates the pivotal roles of rG4s and Ca2+ in promoting Tau aggregation under physiological conditions in vitro, offering insights into potential triggers for tauopathy. [ABSTRACT FROM AUTHOR]