RESUMEN

The generation of quality data from a single-nucleus profiling experiment requires nuclei to be isolated from tissues in a gentle and efficient manner. Nuclei isolation must be carefully optimized across tissue types to preserve nuclear architecture, prevent nucleic acid degradation, and remove unwanted contaminants. Here, we present an optimized workflow for generating a single-nucleus suspension from ocular tissues of the embryonic chicken that is compatible with various downstream workflows. The described protocol enables the rapid isolation of a high yield of aggregate-free nuclei from the embryonic chicken eye without compromising nucleic acid integrity, and the nuclei suspension is compatible with single-nucleus RNA and ATAC sequencing. We detail several stopping points, either via cryopreservation or fixation, to enhance workflow adaptability. Further, we provide a guide through multiple QC points and demonstrate proof-of-principle using two commercially available kits. Finally, we demonstrate that existing in silico genotyping methods can be adopted to computationally derive biological replicates from a single pool of chicken nuclei, greatly reducing the cost of biological replication and allowing researchers to consider sex as a variable during analysis. Together, this tutorial represents a cost-effective, simple, and effective approach to single-nucleus profiling of embryonic chicken eye tissues and is likely to be easily modified to be compatible with similar tissue types.

Asunto(s)

Núcleo Celular , Pollos , Análisis de la Célula Individual , Animales , Núcleo Celular/metabolismo , Núcleo Celular/genética , Embrión de Pollo , Análisis de la Célula Individual/métodos , Ojo/embriología , Ojo/metabolismo , Criopreservación/métodos , Secuenciación de Inmunoprecipitación de Cromatina/métodos

RESUMEN

Retinal progenitor cells (RPCs) are a multipotent and highly proliferative population that give rise to all retinal cell types during organogenesis. Defining their molecular signature is a key step towards identifying suitable approaches to treat visual impairments. Here, we performed RNA sequencing of whole eyes from Xenopus at three embryonic stages and used differential expression analysis to define the transcriptomic profiles of optic tissues containing proliferating and differentiating RPCs during retinogenesis. Gene Ontology and KEGG pathway analyses showed that genes associated with developmental pathways (including Wnt and Hedgehog signaling) were upregulated during the period of active RPC proliferation in early retinal development (Nieuwkoop Faber st. 24 and 27). Developing eyes had dynamic expression profiles and shifted to enrichment for metabolic processes and phototransduction during RPC progeny specification and differentiation (st. 35). Furthermore, conserved adult eye regeneration genes were also expressed during early retinal development, including sox2, pax6, nrl, and Notch signaling components. The eye transcriptomic profiles presented here span RPC proliferation to retinogenesis and include regrowth-competent stages. Thus, our dataset provides a rich resource to uncover molecular regulators of RPC activity and will allow future studies to address regulators of RPC proliferation during eye repair and regrowth.

Asunto(s)

Perfilación de la Expresión Génica , Regulación del Desarrollo de la Expresión Génica , Transcriptoma , Xenopus laevis , Animales , Xenopus laevis/genética , Xenopus laevis/embriología , Transcriptoma/genética , Ojo/metabolismo , Ojo/embriología , Retina/metabolismo , Retina/crecimiento & desarrollo , Diferenciación Celular/genética , Proliferación Celular/genética , Organogénesis/genética , Células Madre/metabolismo , Células Madre/citología , Proteínas de Xenopus/genética , Proteínas de Xenopus/metabolismo , Transducción de Señal/genética

RESUMEN

Biodegradable plastics have been commonly developed and applied as an alternative to traditional plastics, which cause environmental plastic pollution. However, biodegradable plastics still present limitations such as stringent degradation conditions and slow degradation rate, and may cause harm to the environment and organisms. Consequently, in this study, zebrafish was used to evaluate the effects of five biodegradable microplastics (MPs), polyglycolic acid (PGA), polylactic acid (PLA), polybutylene succinate (PBS), polyhydroxyalkanoate (PHA) and polybutylene adipate terephthalate (PBAT) exposure on the early development, retina morphology, visually-mediated behavior, and thyroid signaling at concentrations of 1 mg/L and 100 mg/L. The results indicated that all MPs induced decreased survival rate, reduced body length, smaller eyes, and smaller heads, affecting the early development of zebrafish larvae. Moreover, the thickness of retinal layers, including inner plexiform layer (IPL), outer nuclear layer (ONL), and retinal ganglion layer (RGL) was decreased, and the expression of key genes related to eye and retinal development was abnormally altered after all MPs exposure. Exposure to PBS and PBAT led to abnormal visually-mediated behavior, indicating likely affected the visual function. All MPs could also cause thyroid system disorders, among which alterations in the thyroid hormone receptors (TRs) genes could affect the retinal development of zebrafish larvae. In summary, biodegradable MPs exhibited eye developmental toxicity and likely impaired the visual function in zebrafish larvae. This provided new evidence for revealing the effects of biodegradable plastics on aquatic organism development and environmental risks to aquatic ecosystems.

Asunto(s)

Larva , Glándula Tiroides , Pez Cebra , Animales , Pez Cebra/embriología , Larva/efectos de los fármacos , Larva/crecimiento & desarrollo , Larva/metabolismo , Glándula Tiroides/efectos de los fármacos , Glándula Tiroides/metabolismo , Ojo/efectos de los fármacos , Ojo/crecimiento & desarrollo , Ojo/embriología , Ojo/metabolismo , Contaminantes Químicos del Agua/toxicidad , Plásticos Biodegradables/toxicidad , Sistema Hipotálamo-Hipofisario/efectos de los fármacos , Sistema Hipotálamo-Hipofisario/metabolismo , Conducta Animal/efectos de los fármacos , Microplásticos/toxicidad

RESUMEN

Compound eyes formation in decapod crustaceans occurs after the nauplius stage. However, the key genes and regulatory mechanisms of compound eye development during crustacean embryonic development have not yet been clarified. In this study, RNA-seq was used to investigate the gene expression profiles of Neocaridina denticulata sinensis from nauplius to zoea stage. Based on RNA-seq data analysis, the phototransduction and insect hormone biosynthesis pathways were enriched, and molting-related neuropeptides were highly expressed. There was strong cell proliferation in the embryo prior to compound eye development. The formation of the visual system and the hormonal regulation of hatching were the dominant biological events during compound eye development. The functional analysis of DEGs across all four developmental stages showed that cuticle formation, muscle growth and the establishment of immune system occurred from nauplius to zoea stage. Key genes related to eye development were discovered, including those involved in the determination and differentiation of the eye field, eye-color formation, and visual signal transduction. In conclusion, the results increase the understanding of the molecular mechanism of eye formation in crustacean embryonic stage.

Asunto(s)

Ojo Compuesto de los Artrópodos , Perfilación de la Expresión Génica , Animales , Ojo Compuesto de los Artrópodos/metabolismo , Ojo Compuesto de los Artrópodos/crecimiento & desarrollo , Transcriptoma , Regulación del Desarrollo de la Expresión Génica , Decápodos/genética , Decápodos/crecimiento & desarrollo , Ojo/metabolismo , Ojo/embriología , Ojo/crecimiento & desarrollo

RESUMEN

The conserved bazooka (baz/par3) gene acts as a key regulator of asymmetrical cell divisions across the animal kingdom. Associated Par3/Baz-Par6-aPKC protein complexes are also well known for their role in the establishment of apical/basal cell polarity in epithelial cells. Here we define a novel, positive function of Baz/Par3 in the Notch pathway. Using Drosophila wing and eye development, we demonstrate that Baz is required for Notch signaling activity and optimal transcriptional activation of Notch target genes. Baz appears to act independently of aPKC in these contexts, as knockdown of aPKC does not cause Notch loss-of-function phenotypes. Using transgenic Notch constructs, our data positions Baz activity downstream of activating Notch cleavage steps and upstream of Su(H)/CSL transcription factor complex activity on Notch target genes. We demonstrate a biochemical interaction between NICD and Baz, suggesting that Baz is required for NICD activity before NICD binds to Su(H). Taken together, our data define a novel role of the polarity protein Baz/Par3, as a positive and direct regulator of Notch signaling through its interaction with NICD.

Asunto(s)

Proteínas de Drosophila , Receptores Notch , Transducción de Señal , Alas de Animales , Animales , Proteínas de Drosophila/metabolismo , Proteínas de Drosophila/genética , Receptores Notch/metabolismo , Alas de Animales/metabolismo , Alas de Animales/embriología , Alas de Animales/crecimiento & desarrollo , Regulación del Desarrollo de la Expresión Génica , Unión Proteica , Drosophila melanogaster/metabolismo , Drosophila melanogaster/embriología , Drosophila melanogaster/genética , Ojo/embriología , Ojo/metabolismo , Ojo/crecimiento & desarrollo , Drosophila/metabolismo , Drosophila/embriología , Polaridad Celular , Péptidos y Proteínas de Señalización Intracelular

RESUMEN

The role extracellular matrix (ECM) in multiple events of morphogenesis has been well described, little is known about its specific role in early eye development. One of the first morphogenic events in lens development is placodal thickening, which converts the presumptive lens ectoderm from cuboidal to pseudostratified epithelium. This process occurs in the anterior pre-placodal ectoderm when the optic vesicle approaches the cephalic ectoderm and is regulated by transcription factor Pax6 and secreted BMP4. Since cells and ECM have a dynamic relationship of interdependence and modulation, we hypothesized that the ECM evolves with cell shape changes during lens placode formation. This study investigates changes in optic ECM including both protein distribution deposition, extracellular gelatinase activity and gene expression patterns during early optic development using chicken and mouse models. In particular, the expression of Timp2, a metalloprotease inhibitor, corresponds with a decrease in gelatinase activity within the optic ECM. Furthermore, we demonstrate that optic ECM remodeling depends on BMP signaling in the placode. Together, our findings suggest that the lens placode plays an active role in remodeling the optic ECM during early eye development.

Asunto(s)

Matriz Extracelular , Regulación del Desarrollo de la Expresión Génica , Cristalino , Factor de Transcripción PAX6 , Animales , Matriz Extracelular/metabolismo , Ratones , Cristalino/metabolismo , Cristalino/crecimiento & desarrollo , Cristalino/citología , Factor de Transcripción PAX6/metabolismo , Factor de Transcripción PAX6/genética , Proteínas del Ojo/metabolismo , Proteínas del Ojo/genética , Proteína Morfogenética Ósea 4/metabolismo , Proteína Morfogenética Ósea 4/genética , Embrión de Pollo , Proteínas de Homeodominio/metabolismo , Proteínas de Homeodominio/genética , Inhibidor Tisular de Metaloproteinasa-2/metabolismo , Inhibidor Tisular de Metaloproteinasa-2/genética , Factores de Transcripción Paired Box/metabolismo , Factores de Transcripción Paired Box/genética , Proteínas Represoras/metabolismo , Proteínas Represoras/genética , Transducción de Señal , Pollos/genética , Ojo/metabolismo , Ojo/crecimiento & desarrollo , Ojo/embriología

RESUMEN

Triphenyltin (TPT) is widely used in crop pest control and ship antifouling coatings, which leads to its entry into aquatic environment and poses a threat to aquatic organisms. However, the effects of TPT on the early life stages of wild fish in natural water environments remains unclear. The aim of this study was to assess the toxic effects of TPT on the early life stages of fish under two different environments: field investigation and laboratory experiment. The occurrence of deformities in wild fish embryos and larvae in the Three Gorges Reservoir (TGR) and the developmental toxicity of TPT at different concentrations (0, 0.15, 1.5 and 15 µg Sn/L) to zebrafish embryos and larvae were observed. The results showed that TPT content was higher in wild larvae, reaching 27.21 ng Sn/g w, and the malformation of wild fish larvae mainly occurred in the eyes and spine under natural water environment. Controlled experiment exposure of zebrafish larvae to TPT also resulted in eye and spinal deformities. Gene expression analysis showed that compared with the control group, the expression levels of genes related to eye development (sox2, otx2, stra6 and rx1) and spine development (sox9a and bmp2b) were significantly up-regulated in the 15 µg Sn/L exposure group, which may be the main cause of eye and spine deformity in the early development stage of fish. In addition, the molecular docking results further elucidate that the strong hydrophobic and electrostatic interactions between TPT and protein residues are the main mechanism of TPT induced abnormal gene expression. Based on these results, it can be inferred that TPT is one of the teratogenic factors of abnormal eye and spine development in the early life stage of fish in the TGR. These findings have important implications for understanding the toxicity of TPT on fish.

Asunto(s)

Embrión no Mamífero , Larva , Compuestos Orgánicos de Estaño , Contaminantes Químicos del Agua , Pez Cebra , Animales , Compuestos Orgánicos de Estaño/toxicidad , Contaminantes Químicos del Agua/toxicidad , Larva/efectos de los fármacos , Embrión no Mamífero/efectos de los fármacos , Columna Vertebral/efectos de los fármacos , Columna Vertebral/anomalías , Ojo/efectos de los fármacos , Ojo/embriología

RESUMEN

OBJECTIVE: The aim of this study was to analyze possible alterations (morphological and inflammatory) in the ocular cells of fetuses from mothers with insulin resistance exposed to saturated fatty acids through the period of pregnancy. METHODS: Wistar female rats were induced to develop insulin resistance before pregnancy. Fetuses' skulls were collected on the 20th day of intrauterine life. The rats were separated on the first day of management into two groups according to the diet applied: control group (C): diet containing soybean oil as a source of fat; and saturated fatty acid group (S): diet containing butter as a source of fat. RESULTS: Histological and immunohistochemical analyses have been conducted. The immunohistochemical analyses of interleukin 6, suppressor of cytokine signaling, 3 and signal transducer and activator of transcription 3 did not demonstrate alterations in the expression of proteins in the fetuses of mothers fed with a saturated fatty diet. Moreover, no histopathological changes were noticed between groups. CONCLUSION: The saturated fatty diet does not induce tissue changes or activate the Janus kinase/signal transducer and activator of transcription signaling pathway during eye development in the fetuses of mothers with insulin resistance.

Asunto(s)

Resistencia a la Insulina , Quinasas Janus , Ratas Wistar , Transducción de Señal , Animales , Femenino , Embarazo , Transducción de Señal/efectos de los fármacos , Resistencia a la Insulina/fisiología , Quinasas Janus/metabolismo , Ácidos Grasos/análisis , Grasas de la Dieta/farmacología , Grasas de la Dieta/efectos adversos , Feto/efectos de los fármacos , Inmunohistoquímica , Factor de Transcripción STAT3/metabolismo , Interleucina-6/análisis , Interleucina-6/metabolismo , Ratas , Ojo/embriología , Ojo/efectos de los fármacos

RESUMEN

The intricate steps of human ocular embryology are impacted by cellular and genetic signaling pathways and a myriad of external elements that can affect pregnancy, such as environmental, metabolic, hormonal factors, medications, and intrauterine infections. This review focuses on presenting some of these factors to recognize the multifactorial nature of ocular development and highlight their clinical significance. This review is based on English-language articles sourced from PubMed, Web of Science, and Google Scholar; keywords searched included "ocular development in pregnancy," "ocular embryology," "maternal nutrition," "ophthalmic change," and "visual system development." While some animal models show the disruption of ocular embryology from these external factors, there are limited post-birth assessments in human studies. Much remains unknown about the precise mechanisms of how these external factors can disrupt normal ocular development in utero, and more significant research is needed to understand the pathophysiology of these disruptive effects further. Findings in this review emphasize the importance of additional research in understanding the dynamic association between factors impacting gestation and neonatal ocular development, particularly in the setting of limited resources.

Asunto(s)

Ojo , Exposición Materna , Animales , Femenino , Humanos , Recién Nacido , Embarazo , Ojo/embriología

RESUMEN

Lattice cells (LCs) in the developing Drosophila retina change shape before attaining final form. Previously, we showed that repeated contraction and expansion of apical cell contacts affect these dynamics. Here, we describe another factor, the assembly of a Rho1-dependent medioapical actomyosin ring formed by nodes linked by filaments that contract the apical cell area. Cell area contraction alternates with relaxation, generating pulsatile changes in cell area that exert force on neighboring LCs. Moreover, Rho1 signaling is sensitive to mechanical changes, becoming active when tension decreases and cells expand, while the negative regulator RhoGAP71E accumulates when tension increases and cells contract. This results in cycles of cell area contraction and relaxation that are reciprocally synchronized between adjacent LCs. Thus, mechanically sensitive Rho1 signaling controls pulsatile medioapical actomyosin contraction and coordinates cell behavior across the epithelium. Disrupting the kinetics of pulsing can lead to developmental errors, suggesting this process controls cell shape and tissue integrity during epithelial morphogenesis of the retina.

Asunto(s)

Actomiosina , Drosophila , Ojo , Animales , Citoesqueleto de Actina/fisiología , Actomiosina/fisiología , Citocinesis , Drosophila/embriología , Morfogénesis , Ojo/embriología , Proteínas de Unión al GTP rho/fisiología , Proteínas de Drosophila/fisiología , Retina/citología

RESUMEN

The Hippo signaling pathway is widely considered a master regulator of organ growth because of the prominent overgrowth phenotypes caused by experimental manipulation of its activity. Contrary to this model, we show here that removing Hippo transcriptional output did not impair the ability of the mouse liver and Drosophila eyes to grow to their normal size. Moreover, the transcriptional activity of the Hippo pathway effectors Yap/Taz/Yki did not correlate with cell proliferation, and hyperactivation of these effectors induced gene expression programs that did not recapitulate normal development. Concordantly, a functional screen in Drosophila identified several Hippo pathway target genes that were required for ectopic overgrowth but not normal growth. Thus, Hippo signaling does not instruct normal growth, and the Hippo-induced overgrowth phenotypes are caused by the activation of abnormal genetic programs.

Asunto(s)

Drosophila melanogaster , Ojo , Regulación del Desarrollo de la Expresión Génica , Vía de Señalización Hippo , Hígado , Transcripción Genética , Proteínas Coactivadoras Transcripcionales con Motivo de Unión a PDZ , Proteínas Señalizadoras YAP , Animales , Ratones , Drosophila melanogaster/embriología , Drosophila melanogaster/genética , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Ojo/embriología , Vía de Señalización Hippo/genética , Hígado/embriología , Tamaño de los Órganos , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Transactivadores/genética , Proteínas Coactivadoras Transcripcionales con Motivo de Unión a PDZ/metabolismo , Proteínas Señalizadoras YAP/metabolismo

RESUMEN

MT1-MMP/MMP14 belongs to a subgroup of the matrix metalloproteinases family that presents a transmembrane domain, with a cytosolic tail and the catalytic site exposed to the extracellular space. Deficient mice for this enzyme result in early postnatal death and display severe defects in skeletal, muscle and lung development. By using a transgenic line expressing the LacZ reporter under the control of the endogenous Mt1-mmp promoter, we reported a dynamic spatiotemporal expression pattern for Mt1-mmp from early embryonic to perinatal stages during cardiovascular development and brain formation. Thus, Mt1-mmp shows expression in the endocardium of the heart and the truncus arteriosus by E8.5, and is also strongly detected during vascular system development as well as in endothelial cells. In the brain, LacZ reporter expression was detected in the olfactory bulb, the rostral cerebral cortex and the caudal mesencephalic tectum. LacZ-positive cells were observed in neural progenitors of the spinal cord, neural crest cells and the intersomitic region. In the limb, Mt1-mmp expression was restricted to blood vessels, cartilage primordium and muscles. Detection of the enzyme was confirmed by Western blot and immunohistochemical analysis. We suggest novel functions for this metalloproteinase in angiogenesis, endocardial formation and vascularization during organogenesis. Moreover, Mt1-mmp expression revealed that the enzyme may contribute to heart, muscle and brain throughout development.

Asunto(s)

Sistema Cardiovascular/metabolismo , Embrión de Mamíferos/metabolismo , Desarrollo Embrionario , Ojo/metabolismo , Metaloproteinasa 14 de la Matriz/metabolismo , Morfogénesis , Sistema Nervioso/metabolismo , Animales , Sistema Cardiovascular/embriología , Células Cultivadas , Embrión de Mamíferos/citología , Extremidades/embriología , Extremidades/fisiología , Ojo/embriología , Metaloproteinasa 14 de la Matriz/genética , Ratones , Ratones Endogámicos C57BL , Sistema Nervioso/embriología

RESUMEN

PDGFRα signaling is critically important in ocular development. Previous data on PDGFRα lacks an expression map with high spatial and temporal resolution and lineage information. In this study, we aim to present a detailed PDGFRα expression and lineage map from early embryogenesis to adulthood. PDGFRα-CreER; mT/mG reporter mice were analyzed. mEGFP-positive cells contributed to multiple ocular lineages in a spatiotemporally regulated manner. A dynamic PDGFRα expression was identified in corneal stromal cells, lens epithelial cells, lens fiber cells, and retinal astrocytes during the entire period of eye development, while PDGFRα expression in retinal astrocytes from E17.5 onwards and in Müller glial cells was identified within two weeks after birth. By revealing detailed characterization of gene expression and function, we present a comprehensive map of PDGFRα-expressing cells in the eye for a better understanding of PDGFRα signaling's role during eye development.

Asunto(s)

Linaje de la Célula , Ojo/citología , Ojo/embriología , Receptor alfa de Factor de Crecimiento Derivado de Plaquetas/metabolismo , Animales , Animales Recién Nacidos , Linaje de la Célula/genética , Córnea/citología , Córnea/embriología , Embrión de Mamíferos/citología , Ojo/metabolismo , Regulación del Desarrollo de la Expresión Génica , Ratones , ARN Mensajero/genética , ARN Mensajero/metabolismo , Receptor alfa de Factor de Crecimiento Derivado de Plaquetas/genética , Retina/citología

RESUMEN

The Eyes Absent (EYA) transactivator-phosphatase proteins are important contributors to cell-fate determination processes and to the development of multiple organs. The transcriptional regulatory activity as well as the protein tyrosine phosphatase activities of the EYA proteins can independently contribute to proliferation, differentiation, morphogenesis and tissue homeostasis in different contexts. Aberrant EYA levels or activity are associated with numerous syndromic and non-syndromic developmental disorders, as well as cancers. Commensurate with the multiplicity of biochemical activities carried out by the EYA proteins, they impact upon a range of cellular signaling pathways. Here, we provide a broad overview of the roles played by EYA proteins in development, and highlight the molecular signaling pathways known to be linked with EYA-associated organ development and developmental disorders.

Asunto(s)

Anomalías Congénitas/genética , Ojo/metabolismo , Regulación del Desarrollo de la Expresión Génica , Riñón/metabolismo , Proteínas Tirosina Fosfatasas/genética , Transactivadores/genética , Animales , Anomalías Congénitas/embriología , Anomalías Congénitas/metabolismo , Ojo/embriología , Ojo/crecimiento & desarrollo , Predisposición Genética a la Enfermedad/genética , Humanos , Riñón/embriología , Riñón/crecimiento & desarrollo , Mutación , Proteínas Tirosina Fosfatasas/metabolismo , Transactivadores/metabolismo

RESUMEN

The mechanisms regulating nervous system development are still unknown for a wide variety of taxa. In insects and vertebrates, bone morphogenetic protein (BMP) signaling plays a key role in establishing the dorsal-ventral (D-V) axis and limiting the neuroectoderm to one side of that axis, leading to speculation about the conserved evolution of centralized nervous systems. Studies outside of insects and vertebrates show a more diverse picture of what, if any role, BMP signaling plays in neural development across Bilateria. This is especially true in the morphologically diverse Spiralia (≈Lophotrochozoa). Despite several studies of D-V axis formation and neural induction in spiralians, there is no consensus for how these two processes are related, or whether BMP signaling may have played an ancestral role in either process. To determine the function of BMP signaling during early development of the spiralian annelid Capitella teleta, we incubated embryos and larvae in BMP4 protein for different amounts of time. Adding exogenous BMP protein to early-cleaving C. teleta embryos had a striking effect on formation of the brain, eyes, foregut, and ventral midline in a time-dependent manner. However, adding BMP did not block brain or VNC formation or majorly disrupt the D-V axis. We identified three key time windows of BMP activity. 1) BMP treatment around birth of the 3rd-quartet micromeres caused the loss of the eyes, radialization of the brain, and a reduction of the foregut, which we interpret as a loss of A- and C-quadrant identities with a possible trans-fate switch to a D-quadrant identity. 2) Treatment after the birth of micromere 4d induced formation of a third ectopic brain lobe, eye, and foregut lobe, which we interpret as a trans-fate switch of B-quadrant micromeres to a C-quadrant identity. 3) Continuous BMP treatment from late cleavage (4d â€‹+ â€‹12 â€‹h) through mid-larval stages resulted in a modest expansion of Ct-chrdl expression in the dorsal ectoderm and a concomitant loss of the ventral midline (neurotroch ciliary band). Loss of the ventral midline was accompanied by a collapse of the bilaterally-symmetric ventral nerve cord, although the total amount of neural tissue was not greatly affected. Our results compared with those from other annelids and molluscs suggest that BMP signaling was not ancestrally involved in delimiting neural tissue to one region of the D-V axis. However, the effects of ectopic BMP on quadrant-identity during cleavage stages may represent a non-axial organizing signal that was present in the last common ancestor of annelids and mollusks. Furthermore, in the last common ancestor of annelids, BMP signaling may have functioned in patterning ectodermal fates along the D-V axis in the trunk. Ultimately, studies on a wider range of spiralian taxa are needed to determine the role of BMP signaling during neural induction and neural patterning in the last common ancestor of this group. Ultimately, these comparisons will give us insight into the evolutionary origins of centralized nervous systems and body plans.

Asunto(s)

Proteína Morfogenética Ósea 4/farmacología , Proteínas Morfogenéticas Óseas/metabolismo , Poliquetos/embriología , Poliquetos/metabolismo , Proteínas de Pez Cebra/farmacología , Animales , Tipificación del Cuerpo/efectos de los fármacos , Proteínas Morfogenéticas Óseas/genética , Encéfalo/embriología , Sistema Digestivo/embriología , Embrión no Mamífero/metabolismo , Desarrollo Embrionario , Ojo/embriología , Proteínas del Tejido Nervioso/metabolismo , Sistema Nervioso/embriología , Poliquetos/efectos de los fármacos , Poliquetos/crecimiento & desarrollo , Proteínas Recombinantes/farmacología , Transducción de Señal , Proteína Smad1/genética , Proteína Smad1/metabolismo , Proteína Smad5/genética , Proteína Smad5/metabolismo , Proteína Smad8/genética , Proteína Smad8/metabolismo

RESUMEN

Ocular morphogenesis in vertebrates is a highly organized process, orchestrated largely by intrinsic genetic programs that exhibit stringent spatiotemporal control. Alternations in these genetic instructions can lead to hereditary or nonhereditary congenital disorders, a major cause of childhood visual impairment, and contribute to common late-onset blinding diseases. Currently, limited treatment options exist for clinical phenotypes involving eye development. This review summarizes recent advances in our understanding of early-onset ocular disorders and highlights genetic complexities in development and diseases, specifically focusing on coloboma, congenital glaucoma and Leber congenital amaurosis. We also discuss innovative paradigms for potential therapeutic modalities.

Asunto(s)

Enfermedades Hereditarias del Ojo/genética , Niño , Ojo/embriología , Ojo/metabolismo , Enfermedades Hereditarias del Ojo/patología , Enfermedades Hereditarias del Ojo/terapia , Terapia Genética/métodos , Humanos , Terapia Molecular Dirigida/métodos , Trasplante de Células Madre/métodos

RESUMEN

During development, gene expression is tightly controlled to facilitate the generation of the diverse cell types that form the central nervous system. Brahma-related gene 1 (Brg1, also known as Smarca4) is the catalytic subunit of the SWItch/sucrose nonfermentable (SWI/SNF) chromatin remodeling complex that regulates transcription. We investigated the role of Brg1 between embryonic day 6.5 (E6.5) and E14.5 in Sox2-positive neural stem cells (NSCs). Being without major consequences at E6.5 and E14.5, loss of Brg1 between E7.5 and E12.5 resulted in the formation of rosette-like structures in the subventricular zone, as well as morphological alterations and enlargement of neural retina (NR). Additionally, Brg1-deficient cells showed decreased survival in vitro and in vivo. Furthermore, we uncovered distinct changes in gene expression upon Brg1 loss, pointing towards impaired neuron functions, especially those involving synaptic communication and altered composition of the extracellular matrix. Comparison with mice deficient for integrase interactor 1 (Ini1, also known as Smarcb1) revealed that the enlarged NR was Brg1 specific and was not caused by a general dysfunction of the SWI/SNF complex. These results suggest a crucial role for Brg1 in NSCs during brain and eye development.

Asunto(s)

Encéfalo/embriología , ADN Helicasas/genética , Ojo/embriología , Regulación del Desarrollo de la Expresión Génica/genética , Proteínas Nucleares/genética , Proteína SMARCB1/genética , Factores de Transcripción/genética , Animales , Apoptosis/genética , ADN Helicasas/metabolismo , Matriz Extracelular/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Células-Madre Neurales/citología , Proteínas Nucleares/metabolismo , Factores de Transcripción/metabolismo

RESUMEN

The forkhead transcription factor FOXE3 is critical for vertebrate eye development. Recessive and dominant variants cause human ocular disease but the full range of phenotypes and mechanisms of action for the two classes of variants are unknown. We identified FOXE3 variants in individuals with congenital eye malformations and carried out in vitro functional analysis on selected alleles. Sixteen new recessive and dominant families, including six novel variants, were identified. Analysis of new and previously reported genetic and clinical data demonstrated a broad phenotypic range with an overlap between recessive and dominant disease. Most families with recessive alleles, composed of truncating and forkhead-domain missense variants, had severe corneal opacity (90%; sclerocornea in 47%), aphakia (83%) and microphthalmia (80%), but some had milder features including isolated cataract. The phenotype was most variable for recessive missense variants, suggesting that the functional consequences may be highly dependent on the type of amino acid substitution and its position. When assessed, aniridia or iris hypoplasia were noted in 89% and optic nerve anomalies in 60% of recessive cases, indicating that these defects are also common and may be underrecognized. In dominant pedigrees, caused by extension variants, normal eye size (96%), cataracts (99%) and variable anterior segment anomalies were seen in most, but some individuals had microphthalmia, aphakia or sclerocornea, more typical of recessive disease. Functional studies identified variable effects on the protein stability, DNA binding, nuclear localization and transcriptional activity for recessive FOXE3 variants, whereas dominant alleles showed severe impairment in all areas and dominant-negative characteristics.

Asunto(s)

Anomalías del Ojo/genética , Ojo/embriología , Factores de Transcripción Forkhead/genética , Adolescente , Alelos , Catarata/genética , Niño , Opacidad de la Córnea/genética , Discapacidades del Desarrollo/genética , Ojo/crecimiento & desarrollo , Anomalías del Ojo/enzimología , Femenino , Factores de Transcripción Forkhead/metabolismo , Humanos , Masculino , Mutación , Linaje , Fenotipo

RESUMEN

The basic structure of the eye, which is crucial for visual function, is established during the embryonic process of optic cup morphogenesis. Molecular pathways of specification and patterning are integrated with spatially distinct cell and tissue shape changes to generate the eye, with discrete domains and structural features: retina and retinal pigment epithelium enwrap the lens, and the optic fissure occupies the ventral surface of the eye and optic stalk. Interest in the underlying cell biology of eye morphogenesis has led to a growing body of work, combining molecular genetics and imaging to quantify cellular processes such as adhesion and actomyosin activity. These studies reveal that intrinsic machinery and spatiotemporally specific extrinsic inputs collaborate to control dynamics of cell movements and morphologies. Here we consider recent advances in our understanding of eye morphogenesis, with a focus on the mechanics of eye formation throughout vertebrate systems, including insights and potential opportunities using organoids, which may provide a tractable system to test hypotheses from embryonic models.

Asunto(s)

Ojo/embriología , Disco Óptico/embriología , Actomiosina/metabolismo , Animales , Movimiento Celular , Ojo/metabolismo , Ojo/patología , Humanos , Cristalino/embriología , Morfogénesis/genética , Morfogénesis/fisiología , Disco Óptico/metabolismo , Organogénesis/genética , Organogénesis/fisiología , Retina/embriología , Epitelio Pigmentado de la Retina/citología , Transducción de Señal , Vertebrados/fisiología

RESUMEN

We clarified the properties of visual opsin genes in the marbled sole (Pseudopleuronectes yokohamae) by cDNA sequencing, quantification of the opsin gene expression from the larval to the juvenile stage, and measurement of the maximum absorption spectra (λmax) using photopigment reconstitution. In the marbled sole eye, at least eight visual opsin genes, lws, rh2-a, rh2-b, rh2-c, sws2a, sws2b, sws1, and rh1, were expressed. Quantitative RT-PCR analysis revealed that the expression of opsin genes increased (lws, rh2-c, sws2a, and rh1) or decreased (rh2-a, rh2-b, sws2b, and sws1) from the larval to the juvenile stage. Notably, rh2-a expression was observed only in pre- to mid-metamorphic stage larvae and disappeared after metamorphosis. Thus, pre-metamorphism-specific expression of rh2-a in the marbled sole suggests that its function is restricted to the developmental stage. The reconstituted RH2-A opsin λmax was 470 nm, which is typical of acanthopterygian species. These results strongly suggest that mid-wavelength-sensitive rh2-a expression was diminished drastically in the marbled sole, probably resulting in a shift of spectral sensitivity during its metamorphosis from the larval to the juvenile stage.

Asunto(s)

Lenguado/genética , Rodopsina/genética , Animales , Ojo/embriología , Ojo/metabolismo , Lenguado/embriología , Larva/genética , Larva/crecimiento & desarrollo , Filogenia , ARN Mensajero/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Espectrofotometría , Transcriptoma