RESUMO
How tissues acquire complex shapes is a fundamental question in biology and regenerative medicine. Zebrafish semicircular canals form from invaginations in the otic epithelium (buds) that extend and fuse to form the hubs of each canal. We find that conventional actomyosin-driven behaviors are not required. Instead, local secretion of hyaluronan, made by the enzymes uridine 5'-diphosphate dehydrogenase (ugdh) and hyaluronan synthase 3 (has3), drives canal morphogenesis. Charged hyaluronate polymers osmotically swell with water and generate isotropic extracellular pressure to deform the overlying epithelium into buds. The mechanical anisotropy needed to shape buds into tubes is conferred by a polarized distribution of actomyosin and E-cadherin-rich membrane tethers, which we term cytocinches. Most work on tissue morphogenesis ascribes actomyosin contractility as the driving force, while the extracellular matrix shapes tissues through differential stiffness. Our work inverts this expectation. Hyaluronate pressure shaped by anisotropic tissue stiffness may be a widespread mechanism for powering morphological change in organogenesis and tissue engineering.
Assuntos
Espaço Extracelular/química , Ácido Hialurônico/farmacologia , Morfogênese , Especificidade de Órgãos , Pressão , Canais Semicirculares/citologia , Canais Semicirculares/embriologia , Actomiosina/metabolismo , Animais , Anisotropia , Comportamento Animal , Matriz Extracelular/metabolismo , Ácido Hialurônico/biossíntese , Modelos Biológicos , Morfogênese/efeitos dos fármacos , Especificidade de Órgãos/efeitos dos fármacos , Pressão Osmótica , Canais Semicirculares/diagnóstico por imagem , Comportamento Estereotipado , Peixe-Zebra/embriologia , Proteínas de Peixe-Zebra/metabolismoRESUMO
The DNA-binding protein REST forms complexes with histone deacetylases (HDACs) to repress neuronal genes in non-neuronal cells. In differentiating neurons, REST is downregulated predominantly by transcriptional silencing. Here we report that post-transcriptional inactivation of REST by alternative splicing is required for hearing in humans and mice. We show that, in the mechanosensory hair cells of the mouse ear, regulated alternative splicing of a frameshift-causing exon into the Rest mRNA is essential for the derepression of many neuronal genes. Heterozygous deletion of this alternative exon of mouse Rest causes hair cell degeneration and deafness, and the HDAC inhibitor SAHA (Vorinostat) rescues the hearing of these mice. In humans, inhibition of the frameshifting splicing event by a novel REST variant is associated with dominantly inherited deafness. Our data reveal the necessity for alternative splicing-dependent regulation of REST in hair cells, and they identify a potential treatment for a group of hereditary deafness cases.
Assuntos
Surdez/genética , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Processamento Alternativo/genética , Animais , Linhagem Celular , Éxons , Regulação da Expressão Gênica/genética , Células HEK293 , Células Ciliadas Auditivas/fisiologia , Audição/genética , Audição/fisiologia , Inibidores de Histona Desacetilases/metabolismo , Histona Desacetilases/metabolismo , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Neurônios , Splicing de RNA/genética , Proteínas Repressoras/fisiologia , Fatores de Transcrição , Vorinostat/farmacologiaRESUMO
Deafness or hearing deficits are debilitating conditions. They are often caused by loss of sensory hair cells or defects in their function. In contrast to mammals, nonmammalian vertebrates robustly regenerate hair cells after injury. Studying the molecular and cellular basis of nonmammalian vertebrate hair cell regeneration provides valuable insights into developing cures for human deafness. In this review, we discuss the current literature on hair cell regeneration in the context of other models for sensory cell regeneration, such as the retina and the olfactory epithelium. This comparison reveals commonalities with, as well as differences between, the different regenerating systems, which begin to define a cellular and molecular blueprint of regeneration. In addition, we propose how new technical advances can address outstanding questions in the field.
Assuntos
Células-Tronco Adultas/metabolismo , Orelha Interna/metabolismo , Células Ciliadas Auditivas/fisiologia , Mucosa Olfatória/metabolismo , Regeneração/fisiologia , Retina/metabolismo , Animais , Diferenciação Celular/genética , Proliferação de Células/genética , Citocinas/metabolismo , Orelha Interna/citologia , Células Ciliadas Auditivas/citologia , Células Ciliadas Auditivas/metabolismo , Humanos , Inflamação/genética , Inflamação/metabolismo , Macrófagos/metabolismo , Regeneração/genética , Retina/citologia , Transdução de Sinais/genética , Transdução de Sinais/fisiologia , Ferimentos e Lesões/genética , Ferimentos e Lesões/metabolismoRESUMO
Death of mechanosensory hair cells in the inner ear is a common cause of auditory and vestibular impairment in mammals, which have a limited ability to regrow these cells after damage. In contrast, non-mammalian vertebrates, including zebrafish, can robustly regenerate hair cells after severe organ damage. The zebrafish inner ear provides an understudied model system for understanding hair cell regeneration in organs that are highly conserved with their mammalian counterparts. Here, we quantitatively examine hair cell addition during growth and regeneration of the larval zebrafish inner ear. We used a genetically encoded ablation method to induce hair cell death and we observed gradual regeneration with correct spatial patterning over a 2-week period following ablation. Supporting cells, which surround and are a source of new hair cells, divide in response to hair cell ablation, expanding the possible progenitor pool. In parallel, nascent hair cells arise from direct transdifferentiation of progenitor pool cells temporally uncoupled from supporting cell division. These findings reveal a previously unrecognized mechanism of hair cell regeneration with implications for how hair cells may be encouraged to regenerate in the mammalian ear.
Assuntos
Transdiferenciação Celular , Orelha Interna , Células Ciliadas Auditivas , Regeneração , Células-Tronco , Peixe-Zebra , Animais , Regeneração/fisiologia , Orelha Interna/citologia , Células-Tronco/citologia , Células Ciliadas Auditivas/citologia , Células Ciliadas Auditivas/fisiologia , Proteínas de Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/genética , Animais Geneticamente Modificados , Larva/citologiaRESUMO
Following up on our previous observation that early B cell factor (EBF) sites are enriched in open chromatin of the developing sensory epithelium of the mouse cochlea, we investigated the effect of deletion of Ebf1 on inner ear development. We used a Cre driver to delete Ebf1 at the otocyst stage before development of the cochlea. We examined the cochlea at postnatal day (P) 1 and found that the sensory epithelium had doubled in size but the length of the cochlear duct was unaffected. We also found that deletion of Ebf1 led to ectopic sensory patches in the Kölliker's organ. Innervation of the developing organ of Corti was disrupted with no obvious spiral bundles. The ectopic patches were also innervated. All the extra hair cells (HCs) within the sensory epithelium and Kölliker's organ contained mechanoelectrical transduction channels, as indicated by rapid uptake of FM1-43. The excessive numbers of HCs were still present in the adult Ebf1 conditional knockout (cKO) animal. The animals had significantly elevated auditory brainstem response thresholds, suggesting that this gene is essential for hearing development.
Assuntos
Células Ciliadas Auditivas , Camundongos Knockout , Órgão Espiral , Transativadores , Animais , Transativadores/genética , Transativadores/metabolismo , Órgão Espiral/metabolismo , Células Ciliadas Auditivas/metabolismo , Camundongos , Surdez/genética , Deleção de Genes , Células Labirínticas de Suporte/metabolismo , Cóclea/metabolismo , Potenciais Evocados Auditivos do Tronco EncefálicoRESUMO
The planar polarized organization of hair cells in the vestibular maculae is unique because these sensory organs contain two groups of cells with oppositely oriented stereociliary bundles that meet at a line of polarity reversal (LPR). EMX2 is a transcription factor expressed by one hair cell group that reverses the orientation of their bundles, thereby forming the LPR. We generated Emx2-CreERt2 transgenic mice for genetic lineage tracing and demonstrate Emx2 expression before hair cell specification when the nascent utricle and saccule constitute a continuous prosensory domain. Precursors labeled by Emx2-CreERt2 at this stage give rise to hair cells located along one side of the LPR in the mature utricle or saccule, indicating that this boundary is first established in the prosensory domain. Consistent with this, Emx2-CreERt2 lineage tracing in Dreher mutants, where the utricle and saccule fail to segregate, labels a continuous field of cells along one side of a fused utriculo-saccular-cochlear organ. These observations reveal that LPR positioning is pre-determined in the developing prosensory domain, and that EMX2 expression defines lineages of hair cells with oppositely oriented stereociliary bundles.
Assuntos
Linhagem da Célula , Polaridade Celular , Orelha Interna , Proteínas de Homeodomínio , Fatores de Transcrição , Animais , Camundongos , Linhagem da Célula/genética , Polaridade Celular/genética , Orelha Interna/metabolismo , Orelha Interna/embriologia , Orelha Interna/citologia , Regulação da Expressão Gênica no Desenvolvimento , Células Ciliadas Auditivas/metabolismo , Células Ciliadas Auditivas/citologia , Proteínas de Homeodomínio/metabolismo , Proteínas de Homeodomínio/genética , Camundongos Transgênicos , Sáculo e Utrículo/citologia , Sáculo e Utrículo/metabolismo , Sáculo e Utrículo/embriologia , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genéticaRESUMO
Mechanosensory hair cells of the mature mammalian organ of Corti do not regenerate; consequently, loss of hair cells leads to permanent hearing loss. Although nonmammalian vertebrates can regenerate hair cells from neighboring supporting cells, many humans with severe hearing loss lack both hair cells and supporting cells, with the organ of Corti being replaced by a flat epithelium of nonsensory cells. To determine whether the mature cochlea can produce hair cells in vivo, we reprogrammed nonsensory cells adjacent to the organ of Corti with three hair cell transcription factors: Gfi1, Atoh1, and Pou4f3. We generated numerous hair cell-like cells in nonsensory regions of the cochlea and new hair cells continued to be added over a period of 9 wk. Significantly, cells adjacent to reprogrammed hair cells expressed markers of supporting cells, suggesting that transcription factor reprogramming of nonsensory cochlear cells in adult animals can generate mosaics of sensory cells like those seen in the organ of Corti. Generating such sensory mosaics by reprogramming may represent a potential strategy for hearing restoration in humans.
Assuntos
Surdez , Células Ciliadas Auditivas , Adulto , Animais , Humanos , Fatores de Transcrição/genética , Epitélio , Cóclea , MamíferosRESUMO
Inner ear morphogenesis requires tightly regulated epigenetic and transcriptional control of gene expression. CHD7, an ATP-dependent chromodomain helicase DNA-binding protein, and SOX2, an SRY-related HMG box pioneer transcription factor, are known to contribute to vestibular and auditory system development, but their genetic interactions in the ear have not been explored. Here, we analyzed inner ear development and the transcriptional regulatory landscapes in mice with variable dosages of Chd7 and/or Sox2. We show that combined haploinsufficiency for Chd7 and Sox2 results in reduced otic cell proliferation, severe malformations of semicircular canals, and shortened cochleae with ectopic hair cells. Examination of mice with conditional, inducible Chd7 loss by Sox2CreER reveals a critical period (~E9.5) of susceptibility in the inner ear to combined Chd7 and Sox2 loss. Data from genome-wide RNA-sequencing and CUT&Tag studies in the otocyst show that CHD7 regulates Sox2 expression and acts early in a gene regulatory network to control expression of key otic patterning genes, including Pax2 and Otx2. CHD7 and SOX2 directly bind independently and cooperatively at transcription start sites and enhancers to regulate otic progenitor cell gene expression. Together, our findings reveal essential roles for Chd7 and Sox2 in early inner ear development and may be applicable for syndromic and other forms of hearing or balance disorders.
Assuntos
Redes Reguladoras de Genes , Vestíbulo do Labirinto , Animais , Camundongos , Cóclea , Regulação da Expressão Gênica no Desenvolvimento , Mamíferos , Canais Semicirculares , Fatores de TranscriçãoRESUMO
Developmental senescence is a form of programmed senescence that contributes to morphogenesis during embryonic development. We showed recently that the SIX1 homeoprotein, an essential regulator of organogenesis, is also a repressor of adult cellular senescence. Alterations in the SIX/EYA pathway are linked to the human branchio-oto-renal (BOR) syndrome, a rare congenital disorder associated with defects in the ears, kidneys and branchial arches. Here, we have used Six1-deficient mice, an animal model of the BOR syndrome, to investigate whether dysfunction of senescence underpins the developmental defects associated with SIX1 deficiency. We have focused on the developing inner ear, an organ with physiological developmental senescence that is severely affected in Six1-deficient mice and BOR patients. We show aberrant levels and distribution of senescence markers in Six1-deficient inner ears concomitant with defective morphogenesis of senescent structures. Transcriptomic analysis and ex vivo assays support a link between aberrant senescence and altered morphogenesis in this model, associated with deregulation of the TGFß/BMP pathway. Our results show that misregulation of embryo senescence may lead to genetic developmental disorders, significantly expanding the connection between senescence and disease.
Assuntos
Síndrome Brânquio-Otorrenal , Orelha Interna , Adulto , Humanos , Camundongos , Animais , Proteínas Tirosina Fosfatases/fisiologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas Nucleares/genética , Síndrome Brânquio-Otorrenal/genética , Proteínas de Homeodomínio/metabolismoRESUMO
Our molecular understanding of the early stages of human inner ear development has been limited by the difficulty in accessing fetal samples at early gestational stages. As an alternative, previous studies have shown that inner ear morphogenesis can be partially recapitulated using induced pluripotent stem cells directed to differentiate into inner ear organoids (IEOs). Once validated and benchmarked, these systems could represent unique tools to complement and refine our understanding of human otic differentiation and model developmental defects. Here, we provide the first direct comparisons of the early human embryonic otocyst and fetal sensory organs with human IEOs. We use multiplexed immunostaining and single-cell RNA-sequencing to characterize IEOs at three key developmental steps, providing a new and unique signature of in vitro-derived otic placode, epithelium, neuroblasts and sensory epithelia. In parallel, we evaluate the expression and localization of crucial markers at these equivalent stages in human embryos. Together, our data indicate that the current state-of-the-art protocol enables the specification of bona fide otic tissue, supporting the further application of IEOs to inform inner ear biology and disease.
Assuntos
Orelha Interna , Células-Tronco Pluripotentes , Humanos , Gravidez , Feminino , Epitélio/metabolismo , Diferenciação Celular , OrganoidesRESUMO
Inner ear development requires the coordination of cell types from distinct epithelial, mesenchymal and neuronal lineages. Although we have learned much from animal models, many details about human inner ear development remain elusive. We recently developed an in vitro model of human inner ear organogenesis using pluripotent stem cells in a 3D culture, fostering the growth of a sensorineural circuit, including hair cells and neurons. Despite previously characterizing some cell types, many remain undefined. This study aimed to chart the in vitro development timeline of the inner ear organoid to understand the mechanisms at play. Using single-cell RNA sequencing at ten stages during the first 36â days of differentiation, we tracked the evolution from pluripotency to various ear cell types after exposure to specific signaling modulators. Our findings showcase gene expression that influences differentiation, identifying a plethora of ectodermal and mesenchymal cell types. We also discern aspects of the organoid model consistent with in vivo development, while highlighting potential discrepancies. Our study establishes the Inner Ear Organoid Developmental Atlas (IODA), offering deeper insights into human biology and improving inner ear tissue differentiation.
Assuntos
Orelha Interna , Animais , Humanos , Orelha Interna/metabolismo , Células Ciliadas Auditivas , Organoides , Células Cultivadas , Diferenciação Celular/genéticaRESUMO
Cochlear inner hair cells (IHCs) are primary sound receptors, and are therefore a target for developing treatments for hearing impairment. IHC regeneration in vivo has been widely attempted, although not yet in the IHC-damaged cochlea. Moreover, the extent to which new IHCs resemble wild-type IHCs remains unclear, as is the ability of new IHCs to improve hearing. Here, we have developed an in vivo mouse model wherein wild-type IHCs were pre-damaged and nonsensory supporting cells were transformed into IHCs by ectopically expressing Atoh1 transiently and Tbx2 permanently. Notably, the new IHCs expressed the functional marker vGlut3 and presented similar transcriptomic and electrophysiological properties to wild-type IHCs. Furthermore, the formation efficiency and maturity of new IHCs were higher than those previously reported, although marked hearing improvement was not achieved, at least partly due to defective mechanoelectrical transduction (MET) in new IHCs. Thus, we have successfully regenerated new IHCs resembling wild-type IHCs in many respects in the damaged cochlea. Our findings suggest that the defective MET is a critical barrier that prevents the restoration of hearing capacity and should thus facilitate future IHC regeneration studies.
Assuntos
Células Ciliadas Vestibulares , Perda Auditiva , Camundongos , Animais , Células Ciliadas Auditivas Internas , Cóclea/fisiologia , Perda Auditiva/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genéticaRESUMO
The inner ear sensory epithelia contain mechanosensitive hair cells and supporting cells. Both cell types arise from SOX2-expressing prosensory cells, but the mechanisms underlying the diversification of these cell lineages remain unclear. To determine the transcriptional trajectory of prosensory cells, we established a SOX2-2A-ntdTomato human embryonic stem cell line using CRISPR/Cas9, and performed single-cell RNA-sequencing analyses with SOX2-positive cells isolated from inner ear organoids at various time points between differentiation days 20 and 60. Our pseudotime analysis suggests that vestibular type II hair cells arise primarily from supporting cells, rather than bi-fated prosensory cells in organoids. Moreover, ion channel- and ion-transporter-related gene sets were enriched in supporting cells versus prosensory cells, whereas Wnt signaling-related gene sets were enriched in hair cells versus supporting cells. These findings provide valuable insights into how prosensory cells give rise to hair cells and supporting cells during human inner ear development, and may provide a clue to promote hair cell regeneration from resident supporting cells in individuals with hearing loss or balance disorders.
Assuntos
Células Ciliadas Vestibulares , Vestíbulo do Labirinto , Humanos , Organoides , Células Ciliadas Auditivas , Diferenciação Celular/genéticaRESUMO
Cranial ganglia are aggregates of sensory neurons that mediate distinct types of sensation. The statoacoustic ganglion (SAG) develops into several lobes that are spatially arranged to connect appropriately with hair cells of the inner ear. To investigate the cellular behaviours involved in the 3D organization of the SAG, we use high-resolution confocal imaging of single-cell, labelled zebrafish neuroblasts (NBs), photoconversion, photoablation, and genetic perturbations. We show that otic NBs delaminate out of the otic epithelium in an epithelial-mesenchymal transition-like manner, rearranging apical polarity and primary cilia proteins. We also show that, once delaminated, NBs require RhoGTPases in order to perform active migration. Furthermore, tracking of recently delaminated NBs revealed their directed migration and coalescence around a small population of pioneer SAG neurons. These pioneer SAG neurons, not from otic placode origin, populate the coalescence region before otic neurogenesis begins and their ablation disrupts delaminated NB migratory pathways, consequentially affecting SAG shape. Altogether, this work shows for the first time the role of pioneer SAG neurons in orchestrating SAG development.
Assuntos
Orelha Interna , Peixe-Zebra , Animais , Peixe-Zebra/genética , Diferenciação Celular/genética , Orelha Interna/metabolismo , Células Ciliadas Auditivas/fisiologia , Células Receptoras SensoriaisRESUMO
The auditory organ of Corti is comprised of only two major cell types-the mechanosensory hair cells and their associated supporting cells-both specified from a single pool of prosensory progenitors in the cochlear duct. Here, we show that competence to respond to Atoh1, a transcriptional master regulator necessary and sufficient for induction of mechanosensory hair cells, is established in the prosensory progenitors between E12.0 and 13.5. The transition to the competent state is rapid and is associated with extensive remodeling of the epigenetic landscape controlled by the SoxC group of transcription factors. Conditional loss of Sox4 and Sox11-the two homologous family members transiently expressed in the inner ear at the time of competence establishment-blocks the ability of prosensory progenitors to differentiate as hair cells. Mechanistically, we show that Sox4 binds to and establishes accessibility of early sensory lineage-specific regulatory elements, including ones associated with Atoh1 and its direct downstream targets. Consistent with these observations, overexpression of Sox4 or Sox11 prior to developmental establishment of competence precociously induces hair cell differentiation in the cochlear progenitors. Further, reintroducing Sox4 or Sox11 expression restores the ability of postnatal supporting cells to differentiate as hair cells in vitro and in vivo. Our findings demonstrate the pivotal role of SoxC family members as agents of epigenetic and transcriptional changes necessary for establishing competence for sensory receptor differentiation in the inner ear.
Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos , Fatores de Transcrição SOXC , Animais , Fatores de Transcrição SOXC/genética , Fatores de Transcrição SOXC/metabolismo , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Cóclea/metabolismo , Células Ciliadas Auditivas/metabolismo , Diferenciação Celular , Fatores de Transcrição/metabolismo , Epigênese Genética , Órgão Espiral , Regulação da Expressão Gênica no Desenvolvimento , Mamíferos/metabolismoRESUMO
Type I spiral ganglion neurons (SGNs) are the auditory afferents that transmit sound information from cochlear inner hair cells (IHCs) to the brainstem. These afferents consist of physiological subtypes that differ in their spontaneous firing rate (SR), activation threshold, and dynamic range and have been described as low, medium, and high SR fibers. Lately, single-cell RNA sequencing experiments have revealed three molecularly defined type I SGN subtypes. The extent to which physiological type I SGN subtypes correspond to molecularly defined subtypes is unclear. To address this question, we have generated mouse lines expressing CreERT2 in SGN subtypes that allow for a physiological assessment of molecular subtypes. We show that Lypd1-CreERT2 expressing SGNs represent a well-defined group of neurons that preferentially innervate the IHC modiolar side and exhibit a narrow range of low SRs. In contrast, Calb2-CreERT2 expressing SGNs preferentially innervate the IHC pillar side and exhibit a wider range of SRs, thus suggesting that a strict stratification of all SGNs into three molecular subclasses is not obvious, at least not with the CreERT2 tools used here. Genetically marked neuronal subtypes refine their innervation specificity onto IHCs postnatally during the time when activity is required to refine their molecular phenotype. Type I SGNs thus consist of genetically defined subtypes with distinct physiological properties and innervation patterns. The molecular subtype-specific lines characterized here will provide important tools for investigating the role of the physiologically distinct type I SGNs in encoding sound signals.
Assuntos
Tronco Encefálico , Células Ciliadas Vestibulares , Animais , Camundongos , Cóclea , Células Ciliadas Auditivas Internas , NeurôniosRESUMO
The mouse auditory organ cochlea contains two types of sound receptors: inner hair cells (IHCs) and outer hair cells (OHCs). Tbx2 is expressed in IHCs but repressed in OHCs, and neonatal OHCs that misexpress Tbx2 transdifferentiate into IHC-like cells. However, the extent of this switch from OHCs to IHC-like cells and the underlying molecular mechanism remain poorly understood. Furthermore, whether Tbx2 can transform fully mature adult OHCs into IHC-like cells is unknown. Here, our single-cell transcriptomic analysis revealed that in neonatal OHCs misexpressing Tbx2, 85.6% of IHC genes, including Slc17a8, are upregulated, but only 38.6% of OHC genes, including Ikzf2 and Slc26a5, are downregulated. This suggests that Tbx2 cannot fully reprogram neonatal OHCs into IHCs. Moreover, Tbx2 also failed to completely reprogram cochlear progenitors into IHCs. Lastly, restoring Ikzf2 expression alleviated the abnormalities detected in Tbx2+ OHCs, which supports the notion that Ikzf2 repression by Tbx2 contributes to the transdifferentiation of OHCs into IHC-like cells. Our study evaluates the effects of ectopic Tbx2 expression on OHC lineage development at distinct stages of either male or female mice and provides molecular insights into how Tbx2 disrupts the gene expression profile of OHCs. This research also lays the groundwork for future studies on OHC regeneration.
Assuntos
Células Ciliadas Auditivas Internas , Células Ciliadas Auditivas Externas , Proteínas com Domínio T , Animais , Proteínas com Domínio T/metabolismo , Proteínas com Domínio T/genética , Camundongos , Células Ciliadas Auditivas Internas/metabolismo , Células Ciliadas Auditivas Externas/metabolismo , Feminino , Animais Recém-Nascidos , Transdiferenciação Celular/fisiologia , Transdiferenciação Celular/genética , Masculino , Cóclea/metabolismo , Cóclea/citologia , Camundongos Endogâmicos C57BLRESUMO
The inner ear sensory neurons play a pivotal role in auditory processing and balance control. Though significant progresses have been made, the underlying mechanisms controlling the differentiation and survival of the inner ear sensory neurons remain largely unknown. During development, ISL1 and POU4F transcription factors are co-expressed and are required for terminal differentiation, pathfinding, axon outgrowth and the survival of neurons in the central and peripheral nervous systems. However, little is understood about their functional relationship and regulatory mechanism in neural development. Here, we have knocked out Isl1 or Pou4f1 or both in mice of both sexes. In the absence of Isl1, the differentiation of cochleovestibular ganglion (CVG) neurons is disturbed and with that Isl1-deficient CVG neurons display defects in migration and axon pathfinding. Compound deletion of Isl1 and Pou4f1 causes a delay in CVG differentiation and results in a more severe CVG defect with a loss of nearly all of spiral ganglion neurons (SGNs). Moreover, ISL1 and POU4F1 interact directly in developing CVG neurons and act cooperatively as well as independently in regulating the expression of unique sets of CVG-specific genes crucial for CVG development and survival by binding to the cis-regulatory elements including the promoters of Fgf10, Pou4f2, and Epha5 and enhancers of Eya1 and Ntng2 These findings demonstrate that Isl1 and Pou4f1 are indispensable for CVG development and maintenance by acting epistatically to regulate genes essential for CVG development.
Assuntos
Orelha Interna , Regulação da Expressão Gênica no Desenvolvimento , Animais , Feminino , Masculino , Camundongos , Gânglios/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/genética , Proteínas com Homeodomínio LIM/genética , Proteínas com Homeodomínio LIM/metabolismo , Células Receptoras Sensoriais/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
Early B-cell factor 1 (EBF1) is a basic helix-loop-helix transcription factor essential for the differentiation of various tissues. Our single-cell RNA sequencing data suggest that Ebf1 is expressed in the sensory epithelium of the mouse inner ear. Here, we found that the murine Ebf1 gene and its protein are expressed in the prosensory domain of the inner ear, medial region of the cochlear duct floor, otic mesenchyme, and cochleovestibular ganglion. Ebf1 deletion in mice results in incomplete formation of the spiral limbus and scala tympani, increased number of cells in the organ of Corti and Kölliker's organ, and aberrant course of the spiral ganglion axons. Ebf1 deletion in the mouse cochlear epithelia caused the proliferation of SOX2-positive cochlear cells at E13.5, indicating that EBF1 suppresses the proliferation of the prosensory domain and cells of Kölliker's organ to facilitate the development of appropriate numbers of hair and supporting cells. Furthermore, mice with deletion of cochlear epithelium-specific Ebf1 showed poor postnatal hearing function. Our results suggest that Ebf1 is essential for normal auditory function in mammals.
Assuntos
Orelha Interna , Rampa do Tímpano , Animais , Camundongos , Cóclea/metabolismo , Ducto Coclear , Mamíferos , Gânglio Espiral da Cóclea , Fatores de Transcrição/metabolismoRESUMO
The organ of Corti, located in the cochlea within the inner ear is the receptor organ for hearing. It converts auditory signals into neuronal action potentials that are transmitted to the brain for further processing. The mature organ of Corti consists of a variety of highly differentiated sensory cells that fulfil unique tasks in the processing of auditory signals. The actin and microtubule cytoskeleton play essential function in hearing, however so far, more attention has been paid to the role of actin. Microtubules play important roles in maintaining cellular structure and intracellular transport in virtually all eukaryotic cells. Their functions are controlled by interactions with a large variety of microtubule-associated proteins (MAPs) and molecular motors. Current advances show that tubulin posttranslational modifications, as well as tubulin isotypes could play key roles in modulating microtubule properties and functions in cells. These mechanisms could have various effects on the stability and functions of microtubules in the highly specialised cells of the cochlea. Here, we review the current understanding of the role of microtubule-regulating mechanisms in the function of the cochlea and their implications for hearing, which highlights the importance of microtubules in the field of hearing research.