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[This corrects the article DOI: 10.1371/journal.pgen.1008841.].
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Hypomyelination, a neurological condition characterized by decreased production of myelin sheets by glial cells, often has no known etiology. Elucidating the genetic causes of hypomyelination provides a better understanding of myelination, as well as means to diagnose, council, and treat patients. Here, we present evidence that YIPPEE LIKE 3 (YPEL3), a gene whose developmental role was previously unknown, is required for central and peripheral glial cell development. We identified a child with a constellation of clinical features including cerebral hypomyelination, abnormal peripheral nerve conduction, hypotonia, areflexia, and hypertrophic peripheral nerves. Exome and genome sequencing revealed a de novo mutation that creates a frameshift in the open reading frame of YPEL3, leading to an early stop codon. We used zebrafish as a model system to validate that YPEL3 mutations are causative of neuropathy. We found that ypel3 is expressed in the zebrafish central and peripheral nervous system. Using CRISPR/Cas9 technology, we created zebrafish mutants carrying a genomic lesion similar to that of the patient. Our analysis revealed that Ypel3 is required for development of oligodendrocyte precursor cells, timely exit of the perineurial glial precursors from the central nervous system (CNS), formation of the perineurium, and Schwann cell maturation. Consistent with these observations, zebrafish ypel3 mutants have metabolomic signatures characteristic of oligodendrocyte and Schwann cell differentiation defects, show decreased levels of Myelin basic protein in the central and peripheral nervous system, and develop defasciculated peripheral nerves. Locomotion defects were observed in adult zebrafish ypel3 mutants. These studies demonstrate that Ypel3 is a novel gene required for perineurial cell development and glial myelination.
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Regulación del Desarrollo de la Expresión Génica , Enfermedades Desmielinizantes del Sistema Nervioso Central Hereditarias/genética , Vaina de Mielina/patología , Neurogénesis/genética , Proteínas Supresoras de Tumor/genética , Animales , Plexo Braquial/diagnóstico por imagen , Niño , Análisis Mutacional de ADN , Modelos Animales de Enfermedad , Embrión no Mamífero , Femenino , Mutación del Sistema de Lectura , Sustancia Gris/diagnóstico por imagen , Enfermedades Desmielinizantes del Sistema Nervioso Central Hereditarias/diagnóstico por imagen , Enfermedades Desmielinizantes del Sistema Nervioso Central Hereditarias/patología , Humanos , Imagen por Resonancia Magnética , Neuroglía/patología , Oligodendroglía , Nervio Ciático/diagnóstico por imagen , Sustancia Blanca/diagnóstico por imagen , Secuenciación del Exoma , Pez Cebra , Proteínas de Pez Cebra/genéticaRESUMEN
PURPOSE: Growth differentiation factor 11 (GDF11) is a key signaling protein required for proper development of many organ systems. Only one prior study has associated an inherited GDF11 variant with a dominant human disease in a family with variable craniofacial and vertebral abnormalities. Here, we expand the phenotypic spectrum associated with GDF11 variants and document the nature of the variants. METHODS: We present a cohort of six probands with de novo and inherited nonsense/frameshift (4/6 patients) and missense (2/6) variants in GDF11. We generated gdf11 mutant zebrafish to model loss of gdf11 phenotypes and used an overexpression screen in Drosophila to test variant functionality. RESULTS: Patients with variants in GDF11 presented with craniofacial (5/6), vertebral (5/6), neurological (6/6), visual (4/6), cardiac (3/6), auditory (3/6), and connective tissue abnormalities (3/6). gdf11 mutant zebrafish show craniofacial abnormalities and body segmentation defects that match some patient phenotypes. Expression of the patients' variants in the fly showed that one nonsense variant in GDF11 is a severe loss-of-function (LOF) allele whereas the missense variants in our cohort are partial LOF variants. CONCLUSION: GDF11 is needed for human development, particularly neuronal development, and LOF GDF11 alleles can affect the development of numerous organs and tissues.
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Proteínas Morfogenéticas Óseas , Anomalías Craneofaciales/genética , Factores de Diferenciación de Crecimiento , Animales , Proteínas Morfogenéticas Óseas/genética , Factores de Diferenciación de Crecimiento/genética , Humanos , Mutación Missense , Fenotipo , Columna Vertebral , Pez Cebra/genéticaRESUMEN
People with NR5A1 mutations experience testicular dysgenesis, ovotestes, or adrenal insufficiency, but we do not completely understand the origin of this phenotypic diversity. NR5A1 is expressed in gonadal soma precursor cells before expression of the sex-determining gene SRY. Many fish have two co-orthologs of NR5A1 that likely partitioned ancestral gene subfunctions between them. To explore ancestral roles of NR5A1, we knocked out nr5a1a and nr5a1b in zebrafish. Single-cell RNA-seq identified nr5a1a-expressing cells that co-expressed genes for steroid biosynthesis and the chemokine receptor Cxcl12a in 1-day postfertilization (dpf) embryos, as does the mammalian adrenal-gonadal (interrenal-gonadal) primordium. In 2dpf embryos, nr5a1a was expressed stronger in the interrenal-gonadal primordium than in the early hypothalamus but nr5a1b showed the reverse. Adult Leydig cells expressed both ohnologs and granulosa cells expressed nr5a1a stronger than nr5a1b. Mutants for nr5a1a lacked the interrenal, formed incompletely differentiated testes, had no Leydig cells, and grew far larger than normal fish. Mutants for nr5a1b formed a disorganized interrenal and their gonads completely disappeared. All homozygous mutant genotypes lacked secondary sex characteristics, including male breeding tubercles and female sex papillae, and had exceedingly low levels of estradiol, 11-ketotestosterone, and cortisol. RNA-seq showed that at 21dpf, some animals were developing as females and others were not, independent of nr5a1 genotype. By 35dpf, all mutant genotypes greatly under-expressed ovary-biased genes. Because adult nr5a1a mutants form gonads but lack an interrenal and conversely, adult nr5a1b mutants lack a gonad but have an interrenal, the adrenal, and gonadal functions of the ancestral nr5a1 gene partitioned between ohnologs after the teleost genome duplication, likely owing to reciprocal loss of ancestral tissue-specific regulatory elements. Identifying such elements could provide hints to otherwise unexplained cases of Differences in Sex Development.
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Glándulas Suprarrenales/metabolismo , Proteínas de Unión al ADN/genética , Disgenesia Gonadal/genética , Gónadas/metabolismo , Factores de Transcripción/genética , Proteínas de Pez Cebra/genética , Glándulas Suprarrenales/embriología , Animales , Proteínas de Unión al ADN/metabolismo , Femenino , Gónadas/embriología , Masculino , Fenotipo , Procesos de Determinación del Sexo , Factores de Transcripción/metabolismo , Pez Cebra , Proteínas de Pez Cebra/metabolismoRESUMEN
The semicircular canals in the inner ear sense angular acceleration. In zebrafish, the semicircular canals develop from epithelial projections that grow toward each other and fuse to form pillars. The growth of the epithelial projections is driven by the production and secretion of extracellular matrix components by the epithelium. The conserved oligomeric Golgi 4 protein, Cog4, functions in retrograde vesicle transport within the Golgi and mutations can lead to sensory neural hearing loss. In zebrafish cog4 mutants, the inner ear is smaller and the number of hair cells is reduced. Here, we show that formation of the pillars is delayed and that secretion of extracellular matrix components (ECM) is impaired in cog4-/- mutants. These results show that Cog4 is required for secretion of ECM molecules essential to drive the growth of the epithelial projections and thus regulates morphogenesis of the semicircular canals.
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Epitelio/metabolismo , Canales Semicirculares/crecimiento & desarrollo , Canales Semicirculares/metabolismo , Proteínas de Pez Cebra/metabolismo , Animales , Matriz Extracelular/metabolismo , Regulación del Desarrollo de la Expresión Génica/fisiología , Células Ciliadas Auditivas Internas/metabolismo , Pérdida Auditiva/metabolismo , Morfogénesis/fisiología , Mutación/fisiología , Pez Cebra/crecimiento & desarrollo , Pez Cebra/metabolismoRESUMEN
Morphogenesis and mechanoelectrical transduction of the hair cell mechanoreceptor depend on the correct assembly of Usher syndrome (USH) proteins into highly organized macromolecular complexes. Defects in these proteins lead to deafness and vestibular areflexia in USH patients. Mutations in a non-USH protein, glutaredoxin domain-containing cysteine-rich 1 (GRXCR1), cause non-syndromic sensorineural deafness. To understand the deglutathionylating enzyme function of GRXCR1 in deafness, we generated two grxcr1 zebrafish mutant alleles. We found that hair bundles are thinner in homozygous grxcr1 mutants, similar to the USH1 mutants ush1c (Harmonin) and ush1ga (Sans). In vitro assays showed that glutathionylation promotes the interaction between Ush1c and Ush1ga and that Grxcr1 regulates mechanoreceptor development by preventing physical interaction between these proteins without affecting the assembly of another USH1 protein complex, the Ush1c-Cadherin23-Myosin7aa tripartite complex. By elucidating the molecular mechanism through which Grxcr1 functions, we also identify a mechanism that dynamically regulates the formation of Usher protein complexes.
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Glutarredoxinas/metabolismo , Células Ciliadas Auditivas/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Síndromes de Usher/metabolismo , Proteínas de Pez Cebra/metabolismo , Pez Cebra/metabolismo , Animales , Perros , Glutatión/metabolismo , Aparato de Golgi/metabolismo , Células de Riñón Canino Madin Darby , Actividad Motora , Mutación/genética , Unión Proteica , Transporte de Proteínas , Estereocilios/metabolismo , Especificidad por SustratoRESUMEN
In 2011, the zebrafish research facility at the University of Oregon experienced an outbreak of Mycobacterium marinum that affected both research fish and facility staff. A thorough review of risks to personnel, the zebrafish veterinary care program, and zebrafish husbandry procedures at the research facility followed. In the years since 2011, changes have been implemented throughout the research facility to protect the personnel, the fish colony, and ultimately the continued success of the zebrafish model research program. In this study, we present the history of the outbreak, the changes we implemented, and recommendations to mitigate pathogen outbreaks in zebrafish research facilities.