Biology subjects

Steel, K. P.

Publications and source records attributed to Steel, K. P..

10 recordsLinked to original sources

bioRxiv2026

Zfp719 is a transcription factor important for maintenance of hearing in mice

Zfp719 is a zinc finger transcription factor which, when mutated, results in hearing impairment in mice. Its closest human orthologue, OTK18, has been linked to tinnitus in large human cohorts. Here we present our investigation of the electrophysiological, structural and transcriptional phenotypes in Zfp719tm1a mutant mice. Homozygotes have near-normal hearing at two weeks old, but lose sensitivity rapidly between two and three weeks, suggesting that while Zfp719 is not required for development, it is important for maintaining hearing. Heterozygous mice exhibit progressive hearing impairment for high frequencies at older ages. We observed damaged and degenerating outer hair cells from as early as three weeks old in homozygotes. We carried out bulk RNAseq at three ages and found one gene consistently misregulated, a long non-coding RNA specific to mice, Gm15083. A better understanding of the genes regulated by Zfp719 may shed light on genes and proteins important for maintaining hearing in humans.

neuroscience↗

bioRxiv2026

Estrogen-related receptor gamma is required for normal auditory innervation and is essential for hearing.

Estrogen-related receptor gamma (ESRRG), an orphan nuclear receptor with structural homology to the classical estrogen receptors, is widely recognised as a key metabolic regulator involved in mitochondrial, synaptic, and ion-homeostatic pathways. Previous clinical studies suggest a link between ESRRG and auditory function; for example, ESRRG has been associated with susceptibility to age-related hearing loss in women and implicated in congenital hearing loss. However, the biological mechanisms by which ESRRG may mediate hearing function remain largely unknown. Here, using a combination of in vivo auditory physiological recordings, immunofluorescence analyses, single hair-cell electrophysiology, and transcriptomic approaches, we characterise the phenotype of a new inner-ear conditional Esrrg knockout (Esrrg-cKO) to investigate the role of Esrrg in the auditory system. We found that Esrrg-cKO mice of both sexes develop early-onset hearing loss, as evidenced by elevated auditory brainstem response thresholds and reduced wave 1 amplitudes from two weeks of age. These auditory deficits arise from a combination of early-onset cochlear neuronal and innervation malformations, together with inner hair cell synaptic defects and delayed myelination that persist into adulthood. Furthermore, distortion product otoacoustic emissions and endocochlear potential recordings are normal in Esrrg-cKO mice, and although sensory hair cells are preserved, IHCs retain immature biophysical properties. These findings are consistent with auditory neuropathy, and together with our comparative transcriptome analyses, indicate that Esrrg is an essential molecular driver of normal cochlear innervation and maturation.

neuroscience↗

bioRxiv2025

A non-invasive test for stria vascularis dysfunction

Age-related hearing loss is common in the human population but it is highly heterogeneous in aetiology which has hampered efforts to develop ways of stopping its progression. Three major sites of the initial dysfunction are the sensory hair cells, their innervation, and the stria vascularis which generates the high potassium endolymph maintained at a high endocochlear potential bathing the apical surface of hair cells. Treatments aimed at the initial site-of-lesion may be useful, and diagnostic tools to distinguish the primary site would help stratify clinical trials and facilitate selection of the most suitable treatment for each person. Here we report a new non-invasive test that distinguishes mutant mice with known stria vascularis dysfunction and reduced endocochlear potential from mice with sensory hair cell or neural defects but with normal endocochlear potential. It is based on measuring inter-trial coherence in auditory brainstem responses to individual stimuli. Mice with reduced strial function show good inter-trial coherence compared with the amplitude of the averaged response, while mice with sensory or neural deficits show poor coherence. This method might be useful in humans to predict whether they have reduced strial function, which cannot be measured directly, and identify who would benefit from treatments aimed at boosting strial function.

neuroscience↗

bioRxiv2024

Pathological mechanisms and candidate therapeutic approaches in the hearing loss of mice carrying human MIR96 mutations

Progressive hearing loss is a common problem in the human population with no effective therapeutics currently available. However, it has a strong genetic contribution, and investigating the genes and regulatory interactions underlying hearing loss offers the possibility of identifying therapeutic candidates. Mutations in regulatory genes are particularly useful for this, and an example is the microRNA miR-96, a transcriptional regulator which controls hair cell maturation. Mice and humans carrying mutations in Mir96 all develop hearing loss, but different mutations result in different physiological, structural and transcriptional phenotypes. Here we present our characterisation of two lines of mice carrying different human mutations knocked-in to Mir96. While mice homozygous for either mutation are profoundly deaf from two weeks old, the heterozygous phenotypes differ markedly, with only one mutation resulting in hearing impairment in heterozygosis. Investigations of the structural phenotype showed that one mutation appears to lead to synaptic defects, while the other has a much more severe effect on the hair cell stereociliary bundles. Transcriptome analyses revealed a wide range of misregulated genes in both mutants which were notably dissimilar. We used the transcriptome analyses to investigate candidate therapeutics, and tested one, finding that it delayed the progression of hearing loss in heterozygous mice. Our work adds further support for the importance of the gain of novel targets in microRNA mutants, and offers a proof of concept for the identification of pharmacological interventions to maintain hearing.

genetics↗

bioRxiv2023

Targeted genome editing restores auditory function in adult mice with progressive hearing loss caused by a human microRNA mutation

Mutations in microRNA-96 (MIR96) cause dominant delayed onset hearing loss DFNA50 without treatment. Genome editing has shown efficacy in hearing recovery by intervention in neonatal mice, yet editing in the adult inner ear is necessary for clinical applications. Here, we developed an editing therapy for a C>A point mutation in the seed region of the Mir96 gene, Mir9614C>A associated with hearing loss by screening gRNAs for genome editors and optimizing Cas9 and sgRNA scaffold for efficient and specific mutation editing in vitro. By AAV delivery in pre-symptomatic (3-week-old) and symptomatic (6-week-old) adult Mir9614C>A mutant mice, hair cell on-target editing significantly improved hearing long-term, with an efficacy inversely correlated with injection age. We achieved transient Cas9 expression without the evidence of AAV genomic integration to significantly reduce the safety concerns associated with editing. We developed an AAV-sgmiR96-master system capable of targeting all known human MIR96 mutations. As mouse and human MIR96 sequences share 100% homology, our approach and sgRNA selection for efficient and specific hair cell editing for long-term hearing recovery lays the foundation for future treatment of DFNA50 caused by MIR96 mutations.

bioengineering↗

bioRxiv2023

Reversal of an existing hearing loss in mutant mice

Hearing loss is highly heterogeneous but one common form involves a failure to maintain the local ionic environment of the sensory hair cells reflected in a reduced endocochlear potential. We used a genetic approach to ask if this type of pathology can be reversed, using the Spns2tm1a mouse mutant known to show this defect. By activating Spns2 gene transcription at different ages after the onset of hearing loss we found that an existing auditory impairment can be reversed to give close to normal thresholds for an Auditory Brainstem Response (ABR), at least at low to mid stimulus frequencies. Delaying the activation of Spns2 led to less effective recovery of ABR thresholds suggesting there is a critical period for intervention. Early activation of Spns2 not only led to improvement in auditory function but also to protection of sensory hair cells from secondary degeneration. The genetic approach we have used to establish that this type of hearing loss is in principle reversible could be extended to many other diseases using available mouse resources. Significance statementNeurological diseases are often thought to be irreversible, including hearing loss. In this study we found that one type of hearing loss can be reversed as long as the treatment is delivered within a critical period early in disease progression. This result is a proof-of-concept that hearing loss not only can be avoided but also may be reversed. This genetic approach can be used for a wide range of diseases using existing mouse resources.

genetics↗

bioRxiv2021

Collateral damage: Identification and characterisation of spontaneous mutations causing deafness from a targeted knockout programme

Mice carrying targeted mutations are important for investigating gene function and the role of genes in disease, but the process of culturing embryonic stem cells during the making of a targeted allele offers opportunities for spontaneous mutations to arise. Identifying spontaneous mutations relies on the detection of phenotypes segregating independently of targeted alleles, and many phenotypes are easy to miss if not specifically looked for. Here we present data from a large, targeted knockout programme in which mice were analysed through a phenotyping pipeline. Twenty-five lines out of 1311 displayed different deafness phenotypes that did not segregate with the targeted allele. We have identified 8 different mutations causing deafness in 16 of these 25 lines and characterised the resulting phenotypes. Our data show that spontaneous mutations with observable effects on phenotype are a common side effect of intensive breeding programmes, including those underlying targeted mutation programmes.

genetics↗

bioRxiv2021

Targeted deletion of the RNA-binding protein Caprin1 leads to progressive hearing loss and impairs recovery from noise exposure in mice.

Cell cycle associated protein 1 (Caprin1) is an RNA-binding protein that can regulate the cellular post-transcriptional response to stress. It is a component of both stress granules and neuronal RNA granules and is implicated in neurodegenerative disease, synaptic plasticity and long-term memory formation. Our previous work suggested that Caprin1 also plays a role in the response of the cochlea to stress. Here, targeted inner ear-deletion of Caprin1 in mice leads to an early onset, progressive hearing loss. Auditory brainstem responses from Caprin1-deficient mice show reduced thresholds, with a significant reduction in wave-I amplitudes compared to wildtype. Whilst hair cell structure and numbers were normal, the inner hair cell-spiral ganglion neuron (IHC-SGN) synapse revealed abnormally large post-synaptic GluA2 receptor puncta, a defect consistent with the observed wave-I reduction. Unlike wildtype mice, mild-noise-induced hearing threshold shifts in Caprin1-deficient mice did not recover. Oxidative stress triggered TIA-1/HuR-positive stress granule formation in ex-vivo cochlear explants from Caprin1-deficient mice, showing that stress granules could still be induced. Taken together, these findings suggest that Caprin1 plays a key role in maintenance of auditory function, where it regulates the normal status of the IHC-SGN synapse.

neuroscience↗

bioRxiv2021

Inner hair cell dysfunction in Klhl18 mutant mice leads to low frequency progressive hearing loss

Age-related hearing loss in humans (presbycusis) typically involves impairment of high frequency sensitivity before becoming progressively more severe at lower frequencies. Pathologies initially affecting lower frequency regions of hearing are less common. Here we describe a progressive, predominantly low-frequency hearing impairment in two mutant mouse lines, carrying different mutant alleles of the Klhl18 gene: a spontaneous missense mutation (Klhl18lowf) and a targeted mutation (Klhl18tm1a(KOMP)Wtsi). Both males and females were studied, and the two mutant lines showed similar phenotypes. Auditory brainstem response (ABR) thresholds (a measure of auditory nerve and brainstem neural activity) were normal at 3 weeks old but showed progressive increases from 4 weeks onwards. In contrast, distortion product otoacoustic emission (DPOAE) sensitivity and amplitudes (a reflection of cochlear outer hair cell function) remained normal in mutants. Electrophysiological recordings from the round window of Klhl18lowf mutants at 6 weeks old revealed 1) raised compound action potential thresholds that were similar to ABR thresholds, 2) cochlear microphonic potentials that were normal compared with wildtype and heterozygous control mice and 3) summating potentials that were reduced in amplitude compared to control mice. Scanning electron microscopy showed that Klhl18lowf mutant mice had abnormally tapering inner hair cell stereocilia in the apical half of the cochlea while their synapses appeared normal. These results suggest that Klhl18 is necessary to maintain inner hair cell stereocilia and normal inner hair cell function at low frequencies. Klhl18 mutant mice exhibit an uncommon low frequency hearing impairment with physiological features consistent with Auditory Neuropathy Spectrum Disorder (ANSD). SIGNIFICANCE STATEMENTWe describe a novel progressive hearing loss in Klhl18 mutant mice that affects the lower frequencies of its hearing range. Investigation of two mutant alleles of this gene revealed primary inner hair cell defects affecting the neural output of the cochlea while outer hair cell function appeared normal. The tallest stereocilia of inner hair cells showed an abnormal tapering shape, especially notable in the apical half of the cochlear duct corresponding to the low frequency hearing loss. Our finding of a primary inner hair cell defect associated with raised thresholds for auditory brainstem responses combined with normal outer hair cell function suggests that Klhl18 deficiency and inner hair cell pathology may contribute to Auditory Neuropathy Spectrum Disorder in humans.

neuroscience↗

bioRxiv2019

Hearing impairment due to Mir183/96/182 mutations suggests both loss and gain of function effects

The microRNA miR-96 is important for hearing, as point mutations in humans and mice result in dominant progressive hearing loss. Mir96 is expressed in sensory cells along with Mir182 and Mir183, but the roles of these closely-linked microRNAs are as yet unknown. Here we analyse mice carrying null alleles of Mir182, and of Mir183 and Mir96 together to investigate their roles in hearing. We found that Mir183/96 heterozygous mice had normal hearing and homozygotes were completely deaf with abnormal hair cell stereocilia bundles and reduced numbers of inner hair cell synapses at four weeks old. Mir182 knockout mice developed normal hearing then exhibited progressive hearing loss. Our transcriptional analyses revealed significant changes in a range of other genes, but surprisingly there were fewer genes with altered expression in the organ of Corti of Mir183/96 null mice compared with our previous findings in Mir96Dmdo mutants, which have a point mutation in the miR-96 seed region. This suggests the more severe phenotype of Mir96Dmdo mutants compared with Mir183/96 mutants, including progressive hearing loss in Mir96Dmdo heterozygotes, is likely to be mediated by the gain of novel target genes in addition to the loss of its normal targets. We propose three mechanisms of action of mutant miRNAs; loss of targets that are normally completely repressed, loss of targets whose transcription is normally buffered by the miRNA, and gain of novel targets. Any of these mechanisms could lead to a partial loss of a robust cellular identity and consequent dysfunction.

systems biology↗