Biology subjects

Nicolson, T.

Publications and source records attributed to Nicolson, T..

3 recordsLinked to original sources

bioRxiv2024

Defects in exosome biogenesis are associated with sensorimotor defects in zebrafish vps4a mutants

Mutations in human VPS4A are associated with neurodevelopmental defects, including motor delays and defective muscle tone. VPS4A encodes a AAA-ATPase required for membrane scission, but how mutations in VPS4A lead to impaired control of motor function is not known. Here we identified a mutation in zebrafish vps4a, T248I, that affects sensorimotor transformation. In biochemical experiments we show the T248I mutation reduces the ATPase activity of Vps4a and disassembly of its substrate, ESCRT filaments, which mediate membrane scission. Consistent with the established role for Vps4a in the endocytic pathway and exosome biogenesis, vps4aT248I mutants have enlarged endosomal compartments in the CNS and decreased numbers of circulating exosomes. Resembling the central form of hypotonia in human VPS4A patients, motor neurons and muscle cells are unaffected in mutant zebrafish as touch sensitivity is intact. Unlike somatosensory function, optomotor responses, vestibulospinal (VS), and acoustic startle reflexes are severely impaired in vps4aT248I mutants, indicating a greater sensitivity of these circuits to the T248I mutation. ERG recordings indicate that visual ability is largely reduced in the mutants, however, in vivo imaging of tone-evoked responses in the inner ear and ascending auditory pathway show comparable activity. Further investigation of central pathways in vps4aT248I mutants revealed that sensory cues failed to fully activate neurons in the VS and medial longitudinal fasciculus (MLF) nuclei that directly innervate motor neurons. Our results suggest that a defect in sensorimotor transformation underlies the profound yet selective effects on motor reflexes resulting from the loss of membrane scission mediated by Vps4a. Significance StatementHere we present a T248I mutation in vps4a, which causes sensorimotor defects in zebrafish larvae. Spanning biochemical to systems level analyses, our study indicates that a reduction in Vps4a enzymatic activity leads to endosomal defects and reduced exosome biogenesis, resulting in pronounced deficits in vision and sensorimotor transformation of auditory and vestibular cues. We suggest that the mechanisms underlying this type of dysfunction in zebrafish may also contribute to the condition seen in human patients with de novo mutations in the human VPS4A orthologue.

neuroscience↗

bioRxiv2019

The lhfpl5 ohnologs lhfpl5a and lhfpl5b are required for mechanotransduction in distinct populations of sensory hair cells in zebrafish.

1Hair cells sense and transmit auditory, vestibular, and hydrodynamic information by converting mechanical stimuli into electrical signals. This process of mechano-electrical transduction (MET) requires a mechanically-gated channel localized in the apical stereocilia of hair cells. In mice, lipoma HMGIC fusion partner-like 5 (LHFPL5) acts as an auxiliary subunit of the MET channel whose primary role is to correctly localize PCDH15 and TMC1 to the mechanotransduction complex. Zebrafish have two lhfpl5 genes (lhfpl5a and lhfpl5b), but their individual contributions to MET channel assembly and function have not been analyzed. Here we show that the zebrafish lhfpl5 genes are expressed in discrete populations of hair cells: lhfpl5a expression is restricted to auditory and vestibular hair cells in the inner ear, while lhfpl5b expression is specific to hair cells of the lateral line organ. Consequently, lhfpl5a mutants exhibit defects in auditory and vestibular function, while disruption of lhfpl5b affects hair cells only in the lateral line neuromasts. In contrast to previous reports in mice, localization of Tmc1 does not depend upon Lhfpl5 function in either the inner ear or lateral line organ. In both lhfpl5a and lhfpl5b mutants, GFP-tagged Tmc1 and Tmc2b proteins still localize to the stereocilia of hair cells. Using a stably integrated GFP-Lhfpl5a transgene, we show that the tip link cadherins Pcdh15a and Cdh23, along with the Myo7aa motor protein, are required for correct Lhfpl5a localization at the tips of stereocilia. Our work corroborates the evolutionarily conserved co-dependence between Lhfpl5 and Pcdh15, but also reveals novel requirements for Cdh23 and Myo7aa to correctly localize Lhfpl5a. In addition, our data suggest that targeting of Tmc1 and Tmc2b proteins to stereocilia in zebrafish hair cells occurs independently of Lhfpl5 proteins.

neuroscience↗

bioRxiv2018

Putative pore-forming subunits of the mechano-electrical transduction channel, Tmc1/2b, require Tmie to localize to the site of mechanotransduction in zebrafish sensory hair cells

Mutations in transmembrane inner ear (TMIE) cause deafness in humans; previous studies suggest involvement in the mechano-electrical transduction (MET) complex in sensory hair cells, but TMIEs precise role is unclear. In tmie zebrafish mutants, we observed that GFP-tagged Tmc1 and Tmc2b, which are putative subunits of the MET channel, fail to target to the hair bundle. In contrast, overexpression of Tmie strongly enhances the targeting of Tmc2b-GFP to stereocilia. To identify the motifs of Tmie underlying the regulation of the Tmcs, we systematically deleted or replaced peptide segments. We then assessed localization and functional rescue of each mutated/chimeric form of Tmie in tmie mutants. We determined that the first putative helix was dispensable and identified a novel critical region of Tmie, the extracellular region and transmembrane domain, which mediates both mechanosensitivity and Tmc2b-GFP expression in bundles. Collectively, our results suggest that Tmies role in sensory hair cells is to target and stabilize Tmc subunits to the site of MET.\n\nAuthor summaryHair cells mediate hearing and balance through the activity of a pore-forming channel in the cell membrane. The transmembrane inner ear (TMIE) protein is an essential component of the protein complex that gates this so-called mechanotransduction channel. While it is known that loss of TMIE results in deafness, the function of TMIE within the complex is unclear. Using zebrafish as a deafness model, Pacentine and Nicolson demonstrate that Tmie is required for the localization of other essential complex members, the transmembrane channel-like (Tmc) proteins, Tmc1/2b. They then evaluate twelve unique versions of Tmie, each containing mutations to different domains of Tmie. This analysis reveals that some mutations in Tmie cause dysfunctional gating of the channel as demonstrated through reduced hair cell activity, and that these same dysfunctional versions also display reduced Tmc expression at the normal site of the channel. These findings link hair cell activity with the levels of Tmc in the bundle, reinforcing the currently-debated notion that the Tmcs are the pore-forming subunits of the mechanotransduction channel. The authors conclude that Tmie, through distinct regions, is involved in both trafficking and stabilizing the Tmcs at the site of mechanotransduction.

neuroscience↗