RESUMEN
Alzheimer's disease (AD) is one of the most common forms of neurodegenerative diseases. Apolipoprotein E4 (ApoE4) is the main genetic risk factor in the development of late-onset AD. However, the exact mechanism underlying ApoE4-mediated neurodegeneration remains unclear. We utilized Drosophila melanogaster to examine the neurotoxic effects of various human APOE isoforms when expressed specifically in glial and neural cells. We assessed impacts on mitochondrial dynamics, ER stress, lipid metabolism, and bio-metal ion concentrations in the central nervous system (CNS) of the transgenic flies. Dachshund antibody staining revealed a reduction in the number of Kenyon cells. Behavioral investigations including ethanol tolerance and learning and memory performance demonstrated neuronal dysfunction in APOE4-expressing larvae and adult flies. Transcription level of marf and drp-1 were found to be elevated in APOE4 flies, while atf4, atf6, and xbp-1 s showed down regulation. Enhanced concentrations of triglyceride and cholesterol in the CNS were observed in APOE4 transgenic flies, with especially pronounced effects upon glial-specific expression of the gene. Spectrophotometry of brain homogenate revealed enhanced Fe++ and Zn++ ion levels in conjunction with diminished Cu++ levels upon APOE4 expression. To explore therapeutic strategies, we subjected the flies to heat-shock treatment, aiming to activate heat-shock proteins (HSPs) and assess their potential to mitigate the neurotoxic effects of APOE isoforms. The results showed potential therapeutic benefits for APOE4-expressing flies, hinting at an ability to attenuate memory deterioration. Overall, our findings suggest that APOE4 can alter lipid metabolism, bio metal ion homeostasis, and disrupt the harmonious fission-fusion balance of neuronal and glial mitochondria, ultimately inducing ER stress. These alterations mirror the main clinical manifestations of AD in patients. Therefore, our work underscores the suitability of Drosophila as a fertile model for probing the pathological roles of APOE and furthering our understanding of diverse isoform-specific functions.
Asunto(s)
Enfermedad de Alzheimer , Animales Modificados Genéticamente , Drosophila melanogaster , Animales , Enfermedad de Alzheimer/metabolismo , Enfermedad de Alzheimer/genética , Enfermedad de Alzheimer/patología , Modelos Animales de Enfermedad , Estrés del Retículo Endoplásmico/fisiología , Humanos , Metabolismo de los Lípidos , Apolipoproteínas E/genética , Apolipoproteínas E/metabolismo , Apolipoproteína E4/genética , Apolipoproteína E4/metabolismo , Dinámicas MitocondrialesRESUMEN
BACKGROUND: Neurodevelopmental and intellectual impairments are extremely heterogeneous disorders caused by a diverse variety of genes involved in different molecular pathways and networks. Genetic alterations in cilia, highly-conserved organelles with sensorineural and signal transduction roles can compromise their proper functions and lead to so-called "ciliopathies" featuring intellectual disability (ID) or neurodevelopmental disorders as frequent clinical manifestations. Here, we report several Iranian families affected with ID and other ciliopathy-associated features carrying known and novel variants in two ciliary genes; CEP104 and CEP290. METHODS: Whole exome and Targeted exome sequencing were carried out on affected individuals. Lymphoblastoid cell lines (LCLs) derived from the members of affected families were established for two families carrying CEP104 mutations. RNA and protein expression studies were carried out on these cells using qPCR and Western blot, respectively. RESULTS: A novel homozygous variant; NM_025114.3:c.7341_7344dupACTT p.(Ser2449Thrfs*8) and four previously reported homozygous variants; NM_025114.3:c.322C>T p.(Arg108*), NM_025114.3:c.4393C>T p.(Arg1465*), NM_025114.3:c.5668G>T p.(Gly1890*) and NM_025114.3:c.1666dupA p.(Ile556Asnfs*20) were identified in CEP290. In two other families, two novel homozygous variants; NM_014704:c.2356_2357insTT p.(Cys786Phefs*11) and NM_014704:c.1901_1902insT p.(Leu634Phefs*33) were identified in CEP104, another ciliary gene. qPCR and Western blot analyses showed significantly lower levels of CEP104 transcripts and protein in patients compared to heterozygous or normal family members. CONCLUSION: We emphasize on the clinical variability and pleiotropic phenotypes due to variants of these genes. In conclusion, our findings support the pivotal role of these genes resulting in cognitive and neurodevelopmental features.
Asunto(s)
Discapacidad Intelectual , Antígenos de Neoplasias , Proteínas de Ciclo Celular/genética , Proteínas del Citoesqueleto/genética , Exoma , Humanos , Discapacidad Intelectual/genética , Irán , Mutación , Linaje , Secuenciación del ExomaRESUMEN
Developmental processes require strict regulation of proliferation, differentiation and patterning for the generation of final organ size. Aberrations in these fundamental events are critically important in tumorigenesis and cancer progression. Salt inducible kinases (Siks) are evolutionarily conserved genes involved in diverse biological processes, including salt sensing, metabolism, muscle, cartilage and bone formation, but their role in development remains largely unknown. Recent findings implicate Siks in mitotic control, and in both tumor suppression and progression. Using a tumor model in the Drosophila eye, we show that perturbation of Sik function exacerbates tumor-like tissue overgrowth and metastasis. Furthermore, we show that both Drosophila Sik genes, Sik2 and Sik3, function in eye development processes. We propose that an important target of Siks may be the Notch signaling pathway, as we demonstrate genetic interaction between Siks and Notch pathway members. Finally, we investigate Sik expression in the developing retina and show that Sik2 is expressed in all photoreceptors, basal to cell junctions, while Sik3 appears to be expressed specifically in R3/R4 cells in the developing eye. Combined, our data suggest that Sik genes are important for eye tissue specification and growth, and that their dysregulation may contribute to tumor formation.
Asunto(s)
Drosophila/enzimología , Drosophila/crecimiento & desarrollo , Proteínas Serina-Treonina Quinasas/metabolismo , Receptores Notch/metabolismo , Retina/crecimiento & desarrollo , Animales , Drosophila/metabolismo , Unión Proteica , Retina/metabolismoRESUMEN
The Drosophila inner photoreceptors R7 and R8 are responsible for color vision and their differentiation starts at the third instar larval stage. Only a handful of genes with R7 or R8-cell-specific expression are known. We performed an enhancer-trap screen using a novel piggyBac transposable element, pBGay, carrying a Gal4 sequence under the control of the P promoter to identify novel genes expressed specifically in R7 or R8 cells. From this screen, three lines were analyzed in detail: piggyBacAC109 and piggyBacAC783 are expressed in R8 cells and piggyBacAC887 is expressed in R7 cells at the third instar larval stage and pupal stages. Molecular analysis showed that the piggyBac elements were inserted into the first intron of CG14160 and CG7985 genes and the second intron of unzipped. We show the expression pattern in the developing eye imaginal disc, pupal retina as well as the adult retina. The photoreceptor-specific expression of these genes is reported for the first time and we propose that these lines are useful tools for studying the development of the visual system.
Asunto(s)
Elementos Transponibles de ADN/genética , Proteínas de Drosophila/genética , Drosophila/genética , Células Fotorreceptoras de Invertebrados/metabolismo , Factores de Transcripción/genética , Animales , Diferenciación Celular/genética , Drosophila/crecimiento & desarrollo , Drosophila/metabolismo , Proteínas de Drosophila/metabolismo , Larva/genética , Larva/crecimiento & desarrollo , Larva/metabolismo , Pupa/crecimiento & desarrollo , Pupa/metabolismo , Factores de Transcripción/metabolismoRESUMEN
Target of rapamycin (TOR) is a major signaling pathway and regulator of cell growth. TOR serves as a hub of many signaling routes, and is implicated in the pathophysiology of numerous human diseases, including cancer, diabetes, and neurodegeneration. Therefore, elucidation of unknown components of TOR signaling that could serve as potential biomarkers and drug targets has a great clinical importance. In this study, our aim is to integrate transcriptomics, interactomics, and regulomics data in Saccharomyces cerevisiae using a network-based multiomics approach to enlighten previously unidentified, potential components of TOR signaling. We constructed the TOR-signaling protein interaction network, which was used as a template to search for TOR-mediated rapamycin and caffeine signaling paths. We scored the paths passing from at least one component of TOR Complex 1 or 2 (TORC1/TORC2) using the co-expression levels of the genes in the transcriptome data of the cells grown in the presence of rapamycin or caffeine. The resultant network revealed seven hitherto unannotated proteins, namely, Atg14p, Rim20p, Ret2p, Spt21p, Ylr257wp, Ymr295cp, and Ygr017wp, as potential components of TOR-mediated rapamycin and caffeine signaling in yeast. Among these proteins, we suggest further deciphering of the role of Ylr257wp will be particularly informative in the future because it was the only protein whose removal from the constructed network hindered the signal transduction to the TORC1 effector kinase Npr1p. In conclusion, this study underlines the value of network-based multiomics integrative data analysis in discovering previously unidentified components of the signaling networks by revealing potential components of TOR signaling for future experimental validation.
Asunto(s)
Serina-Treonina Quinasas TOR/metabolismo , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Diana Mecanicista del Complejo 2 de la Rapamicina/metabolismo , Proteómica , Saccharomyces cerevisiae/patogenicidad , Proteínas de Saccharomyces cerevisiae/metabolismo , Transducción de SeñalRESUMEN
The advent of high-throughput sequencing technologies has led to an exponential increase in the identification of novel disease-causing genes in highly heterogeneous diseases. A novel frameshift mutation in CNKSR1 gene was detected by Next-Generation Sequencing (NGS) in an Iranian family with syndromic autosomal recessive intellectual disability (ARID). CNKSR1 encodes a connector enhancer of kinase suppressor of Ras 1, which acts as a scaffold component for receptor tyrosine kinase in mitogen-activated protein kinase (MAPK) cascades. CNKSR1 interacts with proteins which have already been shown to be associated with intellectual disability (ID) in the MAPK signaling pathway and promotes cell migration through RhoA-mediated c-Jun N-terminal kinase (JNK) activation. Lack of CNKSR1 transcripts and protein was observed in lymphoblastoid cells derived from affected patients using qRT-PCR and western blot analysis, respectively. Furthermore, RNAi-mediated knockdown of cnk, the CNKSR1 orthologue in Drosophila melanogaster brain, led to defects in eye and mushroom body (MB) structures. In conclusion, our findings support the possible role of CNKSR1 in brain development which can lead to cognitive impairment.
Asunto(s)
Discapacidad Intelectual/genética , Péptidos y Proteínas de Señalización Intracelular/genética , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Adolescente , Adulto , Animales , Encéfalo/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster , Familia , Femenino , Genes Recesivos , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Discapacidad Intelectual/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Irán , Sistema de Señalización de MAP Quinasas/genética , Masculino , Mutación , Linaje , Transducción de Señal , SíndromeRESUMEN
Exploring genes and pathways underlying intellectual disability (ID) provides insight into brain development and function, clarifying the complex puzzle of how cognition develops. As part of ongoing systematic studies to identify candidate ID genes, linkage analysis and next-generation sequencing revealed Zinc Finger and BTB Domain Containing 11 (ZBTB11) as a novel candidate ID gene. ZBTB11 encodes a little-studied transcription regulator, and the two identified missense variants in this study are predicted to disrupt canonical Zn2+-binding residues of its C2H2 zinc finger domain, leading to possible altered DNA binding. Using HEK293T cells transfected with wild-type and mutant GFP-ZBTB11 constructs, we found the ZBTB11 mutants being excluded from the nucleolus, where the wild-type recombinant protein is predominantly localized. Pathway analysis applied to ChIP-seq data deposited in the ENCODE database supports the localization of ZBTB11 in nucleoli, highlighting associated pathways such as ribosomal RNA synthesis, ribosomal assembly, RNA modification and stress sensing, and provides a direct link between subcellular ZBTB11 location and its function. Furthermore, given the report of prominent brain and spinal cord degeneration in a zebrafish Zbtb11 mutant, we investigated ZBTB11-ortholog knockdown in Drosophila melanogaster brain by targeting RNAi using the UAS/Gal4 system. The observed approximate reduction to a third of the mushroom body size-possibly through neuronal reduction or degeneration-may affect neuronal circuits in the brain that are required for adaptive behavior, specifying the role of this gene in the nervous system. In conclusion, we report two ID families segregating ZBTB11 biallelic mutations disrupting Zn2+-binding motifs and provide functional evidence linking ZBTB11 dysfunction to this phenotype.
Asunto(s)
Discapacidad Intelectual/genética , Sistema Nervioso/metabolismo , Proteínas Represoras/genética , Médula Espinal/metabolismo , Proteínas de Pez Cebra/genética , Animales , Modelos Animales de Enfermedad , Drosophila melanogaster/genética , Regulación de la Expresión Génica , Técnicas de Silenciamiento del Gen , Células HEK293 , Humanos , Discapacidad Intelectual/patología , Mutación Missense/genética , Sistema Nervioso/patología , Fenotipo , Unión Proteica , Médula Espinal/patología , Pez Cebra/genéticaRESUMEN
MOTIVATION: Understanding the regulatory mechanisms governing eye development of the model organism Drosophila melanogaster (D. m.) requires structured knowledge of the involved genes and proteins, their interactions, and dynamic expression patterns. Especially the latter information is however to a large extent scattered throughout the literature. RESULTS: FlyOde is an online platform for the systematic assembly of data on D. m. eye development. It consists of data on eye development obtained from the literature, and a web interface for users to interactively display these data as a gene regulatory network. Our manual curation process provides high standard structured data, following a specifically designed ontology. Visualization of gene interactions provides an overview of network topology, and filtering according to user-defined expression patterns makes it a versatile tool for daily tasks, as demonstrated by usage examples. Users are encouraged to submit additional data via a simple online form.
Asunto(s)
Ansiedad/fisiopatología , Circulación Coronaria/fisiología , Vasos Coronarios/fisiología , Depresión/fisiopatología , Flujo Sanguíneo Regional/fisiología , Angiografía Coronaria , Electrocardiografía , Femenino , Humanos , Masculino , Estudios Prospectivos , Psicometría/métodos , Encuestas y CuestionariosRESUMEN
Genome control is operated by transcription factors (TFs) controlling their target genes by binding to promoters and enhancers. Conceptually, the interactions between TFs, their binding sites, and their functional targets are represented by gene regulatory networks (GRNs). Deciphering in vivo GRNs underlying organ development in an unbiased genome-wide setting involves identifying both functional TF-gene interactions and physical TF-DNA interactions. To reverse engineer the GRNs of eye development in Drosophila, we performed RNA-seq across 72 genetic perturbations and sorted cell types and inferred a coexpression network. Next, we derived direct TF-DNA interactions using computational motif inference, ultimately connecting 241 TFs to 5,632 direct target genes through 24,926 enhancers. Using this network, we found network motifs, cis-regulatory codes, and regulators of eye development. We validate the predicted target regions of Grainyhead by ChIP-seq and identify this factor as a general cofactor in the eye network, being bound to thousands of nucleosome-free regions.
Asunto(s)
Ojo Compuesto de los Artrópodos/metabolismo , Drosophila/genética , Redes Reguladoras de Genes , Motivos de Nucleótidos , Transcriptoma , Animales , Ojo Compuesto de los Artrópodos/crecimiento & desarrollo , Drosophila/crecimiento & desarrollo , Drosophila/metabolismo , Regulación del Desarrollo de la Expresión GénicaRESUMEN
In the Drosophila optic lobes, the medulla processes visual information coming from inner photoreceptors R7 and R8 and from lamina neurons. It contains approximately 40,000 neurons belonging to more than 70 different types. Here we describe how precise temporal patterning of neural progenitors generates these different neural types. Five transcription factors-Homothorax, Eyeless, Sloppy paired, Dichaete and Tailless-are sequentially expressed in a temporal cascade in each of the medulla neuroblasts as they age. Loss of Eyeless, Sloppy paired or Dichaete blocks further progression of the temporal sequence. We provide evidence that this temporal sequence in neuroblasts, together with Notch-dependent binary fate choice, controls the diversification of the neuronal progeny. Although a temporal sequence of transcription factors had been identified in Drosophila embryonic neuroblasts, our work illustrates the generality of this strategy, with different sequences of transcription factors being used in different contexts.
Asunto(s)
Encéfalo/citología , Diferenciación Celular , Linaje de la Célula , Drosophila melanogaster/citología , Células-Madre Neurales/citología , Neuronas/citología , Neuronas/metabolismo , Animales , Encéfalo/crecimiento & desarrollo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/anatomía & histología , Drosophila melanogaster/metabolismo , Femenino , Regulación de la Expresión Génica , Masculino , Células-Madre Neurales/metabolismo , Factores de Tiempo , Factores de Transcripción/metabolismo , Vías Visuales/citologíaRESUMEN
The functionality of sensory neurons is defined by the expression of specific sensory receptor genes. During the development of the Drosophila larval eye, photoreceptor neurons (PRs) make a binary choice to express either the blue-sensitive Rhodopsin 5 (Rh5) or the green-sensitive Rhodopsin 6 (Rh6). Later during metamorphosis, ecdysone signaling induces a cell fate and sensory receptor switch: Rh5-PRs are re-programmed to express Rh6 and become the eyelet, a small group of extraretinal PRs involved in circadian entrainment. However, the genetic and molecular mechanisms of how the binary cell fate decisions are made and switched remain poorly understood. We show that interplay of two transcription factors Senseless (Sens) and Hazy control cell fate decisions, terminal differentiation of the larval eye and its transformation into eyelet. During initial differentiation, a pulse of Sens expression in primary precursors regulates their differentiation into Rh5-PRs and repression of an alternative Rh6-cell fate. Later, during the transformation of the larval eye into the adult eyelet, Sens serves as an anti-apoptotic factor in Rh5-PRs, which helps in promoting survival of Rh5-PRs during metamorphosis and is subsequently required for Rh6 expression. Comparably, during PR differentiation Hazy functions in initiation and maintenance of rhodopsin expression. Hazy represses Sens specifically in the Rh6-PRs, allowing them to die during metamorphosis. Our findings show that the same transcription factors regulate diverse aspects of larval and adult PR development at different stages and in a context-dependent manner.
Asunto(s)
Linaje de la Célula/genética , Proteínas de Drosophila/genética , Ojo/crecimiento & desarrollo , Metamorfosis Biológica/genética , Proteínas Nucleares/genética , Factores de Transcripción/genética , Animales , Diferenciación Celular/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/genética , Drosophila melanogaster/crecimiento & desarrollo , Drosophila melanogaster/metabolismo , Ecdisona/biosíntesis , Ojo/metabolismo , Regulación del Desarrollo de la Expresión Génica , Larva/crecimiento & desarrollo , Larva/metabolismo , Proteínas Nucleares/metabolismo , Células Fotorreceptoras de Invertebrados/citología , Células Fotorreceptoras de Invertebrados/metabolismo , Rodopsina/genética , Rodopsina/metabolismo , Células Receptoras Sensoriales/citología , Células Receptoras Sensoriales/metabolismo , Factores de Transcripción/metabolismoRESUMEN
Sensory systems with high discriminatory power use neurons that express only one of several alternative sensory receptor proteins. This exclusive receptor gene expression restricts the sensitivity spectrum of neurons and is coordinated with the choice of their synaptic targets. However, little is known about how it is maintained throughout the life of a neuron. Here we show that the green-light sensing receptor rhodopsin 6 (Rh6) acts to exclude an alternative blue-sensitive rhodopsin 5 (Rh5) from a subset of Drosophila R8 photoreceptor neurons. Loss of Rh6 leads to a gradual expansion of Rh5 expression into all R8 photoreceptors of the ageing adult retina. The Rh6 feedback signal results in repression of the rh5 promoter and can be mimicked by other Drosophila rhodopsins; it is partly dependent on activation of rhodopsin by light, and relies on G(αq) activity, but not on the subsequent steps of the phototransduction cascade. Our observations reveal a thus far unappreciated spectral plasticity of R8 photoreceptors, and identify rhodopsin feedback as an exclusion mechanism.
Asunto(s)
Drosophila melanogaster/citología , Drosophila melanogaster/metabolismo , Retroalimentación Sensorial , Células Fotorreceptoras de Invertebrados/metabolismo , Rodopsina/metabolismo , Animales , Regulación hacia Abajo , Proteínas de Drosophila/deficiencia , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/genética , Regiones Promotoras Genéticas/genética , Retina/citología , Rodopsina/deficiencia , Rodopsina/genéticaRESUMEN
The Drosophila eye is a mosaic that results from the stochastic distribution of two ommatidial subtypes. Pale and yellow ommatidia can be distinguished by the expression of distinct rhodopsins and other pigments in their inner photoreceptors (R7 and R8), which are implicated in color vision. The pale subtype contains ultraviolet (UV)-absorbing Rh3 in R7 and blue-absorbing Rh5 in R8. The yellow subtype contains UV-absorbing Rh4 in R7 and green-absorbing Rh6 in R8. The exclusive expression of one rhodopsin per photoreceptor is a widespread phenomenon, although exceptions exist. The mechanisms leading to the exclusive expression or to co-expression of sensory receptors are currently not known. We describe a new class of ommatidia that co-express rh3 and rh4 in R7, but maintain normal exclusion between rh5 and rh6 in R8. These ommatidia, which are localized in the dorsal eye, result from the expansion of rh3 into the yellow-R7 subtype. Genes from the Iroquois Complex (Iro-C) are necessary and sufficient to induce co-expression in yR7. Iro-C genes allow photoreceptors to break the "one receptor-one neuron" rule, leading to a novel subtype of broad-spectrum UV- and green-sensitive ommatidia.
Asunto(s)
Proteínas de Drosophila/genética , Drosophila/genética , Proteínas de Homeodominio/genética , Rodopsina/genética , Animales , Células Cultivadas , Drosophila/metabolismo , Proteínas de Drosophila/metabolismo , Expresión Génica , Retina/metabolismo , Rodopsina/metabolismo , Células Receptoras Sensoriales/metabolismoAsunto(s)
Drosophila melanogaster/metabolismo , Ingeniería Genética/métodos , Células Fotorreceptoras de Invertebrados/metabolismo , Retina/metabolismo , Pigmentos Retinianos/genética , Opsinas de Bastones/genética , Animales , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/genética , Drosophila melanogaster/fisiología , Retina/fisiología , Pigmentos Retinianos/metabolismo , Opsinas de Bastones/metabolismoRESUMEN
One of the most fascinating topics in biology is to understand the development of highly differentiated cells such as photoreceptors (PRs). This process involves successive steps, starting with the generation of the eye primordium, recruitment and specification of PRs and finally, expression of the proper rhodopsin, the photopigment that initiates the signaling cascade underlying light input excitation. In this review, we describe the sequential steps that take place in the Drosophila eye, from the initial neuronal specification of PRs through their full maturation, focusing specifically on the transcription factors and signaling pathways involved in controlling the precise expression of different rhodopsins in specialized PRs.
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Proteínas de Drosophila/genética , Drosophila/genética , Regulación del Desarrollo de la Expresión Génica , Células Fotorreceptoras/metabolismo , Animales , Tipificación del Cuerpo/genética , Tipificación del Cuerpo/fisiología , Drosophila/citología , Drosophila/crecimiento & desarrollo , Proteínas de Drosophila/fisiología , Modelos Genéticos , Células Fotorreceptoras/citología , Células Fotorreceptoras/crecimiento & desarrollo , Retina/citología , Retina/crecimiento & desarrollo , Retina/metabolismoAsunto(s)
Neuronas Receptoras Olfatorias/metabolismo , Receptores Notch/metabolismo , Receptores Odorantes/metabolismo , Olfato/fisiología , Animales , Encéfalo/citología , Encéfalo/crecimiento & desarrollo , Encéfalo/metabolismo , Drosophila/citología , Drosophila/crecimiento & desarrollo , Drosophila/metabolismo , Regulación del Desarrollo de la Expresión Génica/genética , Humanos , Vías Olfatorias/citología , Vías Olfatorias/crecimiento & desarrollo , Vías Olfatorias/metabolismo , Neuronas Receptoras Olfatorias/citología , Neuronas Receptoras Olfatorias/crecimiento & desarrollo , Receptores Notch/genéticaRESUMEN
Drosophila colour vision is achieved by R7 and R8 photoreceptor cells present in every ommatidium. The fly retina contains two types of ommatidia, called 'pale' and 'yellow', defined by different rhodopsin pairs expressed in R7 and R8 cells. Similar to the human cone photoreceptors, these ommatidial subtypes are distributed stochastically in the retina. The choice between pale versus yellow ommatidia is made in R7 cells, which then impose their fate onto R8. Here we report that the Drosophila dioxin receptor Spineless is both necessary and sufficient for the formation of the ommatidial mosaic. A short burst of spineless expression at mid-pupation in a large subset of R7 cells precedes rhodopsin expression. In spineless mutants, all R7 and most R8 cells adopt the pale fate, whereas overexpression of spineless is sufficient to induce the yellow R7 fate. Therefore, this study suggests that the entire retinal mosaic required for colour vision is defined by the stochastic expression of a single transcription factor, Spineless.
Asunto(s)
Percepción de Color/fisiología , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/embriología , Drosophila melanogaster/fisiología , Regulación del Desarrollo de la Expresión Génica , Receptores de Hidrocarburo de Aril/metabolismo , Retina/embriología , Retina/fisiología , Animales , Diferenciación Celular , Linaje de la Célula , Color , Proteínas de Drosophila/genética , Drosophila melanogaster/citología , Drosophila melanogaster/genética , Modelos Biológicos , Mutación/genética , Receptores de Hidrocarburo de Aril/genética , Retina/citología , Células Fotorreceptoras Retinianas Conos/citología , Células Fotorreceptoras Retinianas Conos/embriología , Células Fotorreceptoras Retinianas Conos/fisiología , Procesos EstocásticosRESUMEN
Slow Wallerian degeneration (Wld(S)) mutant mice express a chimeric nuclear protein that protects sick or injured axons from degeneration. The C-terminal region, derived from NAD(+) synthesizing enzyme Nmnat1, is reported to confer neuroprotection in vitro. However, an additional role for the N-terminal 70 amino acids (N70), derived from multiubiquitination factor Ube4b, has not been excluded. In wild-type Ube4b, N70 is part of a sequence essential for ubiquitination activity but its role is not understood. We report direct binding of N70 to valosin-containing protein (VCP; p97/Cdc48), a protein with diverse cellular roles including a pivotal role in the ubiquitin proteasome system. Interaction with Wld(S) targets VCP to discrete intranuclear foci where ubiquitin epitopes can also accumulate. Wld(S) lacking its N-terminal 16 amino acids (N16) neither binds nor redistributes VCP, but continues to accumulate in intranuclear foci, targeting its intrinsic NAD(+) synthesis activity to these same foci. Wild-type Ube4b also requires N16 to bind VCP, despite a more C-terminal binding site in invertebrate orthologues. We conclude that N-terminal sequences of Wld(S) protein influence the intranuclear location of both ubiquitin proteasome and NAD(+) synthesis machinery and that an evolutionary recent sequence mediates binding of mammalian Ube4b to VCP.
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Proteínas de Ciclo Celular/metabolismo , Núcleo Celular/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Adenosina Trifosfatasas , Secuencia de Aminoácidos , Animales , Células COS , Proteínas de Ciclo Celular/química , Células Cultivadas , Chlorocebus aethiops , Evolución Molecular , Células HeLa , Humanos , Espacio Intranuclear/metabolismo , Ratones , Ratones Endogámicos C57BL , Datos de Secuencia Molecular , Proteínas del Tejido Nervioso/química , Nicotinamida-Nucleótido Adenililtransferasa/metabolismo , Unión Proteica , Transporte de Proteínas , Ratas , Proteínas Recombinantes de Fusión/metabolismo , Ubiquitina/metabolismo , Proteína que Contiene ValosinaRESUMEN
OBJECTIVE: Determine the effect on prognosis of Langerhans cell infiltration in cancer of the larynx. STUDY DESIGN AND SETTING: Retrospective review of 72 surgically treated patients with T1-3 lesions. The streptavidin-biotin method to determine Langerhans cell infiltration, which was graded as 1+, 2+, and 3+. RESULTS: A higher degree of Langerhans cell infiltration was significantly associated with less cervical lymph node metastasis, longer disease-free survival, less locoregional recurrence, and less clinical N-positivity ( P < 0.05). According to multivariant analysis, Langerhans cell infiltration was independently related to disease-free survival and recurrence ( P < 0.05). CONCLUSIONS: Langerhans cell infiltration is prognostically important in cancer of the larynx. More intense infiltration is significantly related to prolonged disease-free survival, less locoregional recurrence, less cervical lymph node metastasis, and less clinical N-positivity. SIGNIFICANCE: Langerhans cell infiltration may be determined on a biopsy specimen and this information may be useful in deciding about elective neck dissection. Patients with mild infiltration may have a higher risk of locoregional recurrence, shorter disease-free survival, and therefore they suggest a poor prognosis.