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1.
Cell ; 162(6): 1391-403, 2015 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-26359990

RESUMO

How metazoan mechanotransduction channels sense mechanical stimuli is not well understood. The NOMPC channel in the transient receptor potential (TRP) family, a mechanotransduction channel for Drosophila touch sensation and hearing, contains 29 Ankyrin repeats (ARs) that associate with microtubules. These ARs have been postulated to act as a tether that conveys force to the channel. Here, we report that these N-terminal ARs form a cytoplasmic domain essential for NOMPC mechanogating in vitro, mechanosensitivity of touch receptor neurons in vivo, and touch-induced behaviors of Drosophila larvae. Duplicating the ARs elongates the filaments that tether NOMPC to microtubules in mechanosensory neurons. Moreover, microtubule association is required for NOMPC mechanogating. Importantly, transferring the NOMPC ARs to mechanoinsensitive voltage-gated potassium channels confers mechanosensitivity to the chimeric channels. These experiments strongly support a tether mechanism of mechanogating for the NOMPC channel, providing insights into the basis of mechanosensitivity of mechanotransduction channels.


Assuntos
Proteínas de Drosophila/química , Proteínas de Drosophila/metabolismo , Drosophila/metabolismo , Mecanotransdução Celular , Canais de Potencial de Receptor Transitório/química , Canais de Potencial de Receptor Transitório/metabolismo , Animais , Drosophila/citologia , Drosophila/crescimento & desenvolvimento , Canal de Potássio Kv1.2/metabolismo , Larva/citologia , Larva/metabolismo , Microtúbulos/metabolismo , Estrutura Terciária de Proteína , Tato
2.
Bioessays ; 46(5): e2300240, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38593308

RESUMO

The compound eyes of insects exhibit stunning variation in size, structure, and function, which has allowed these animals to use their vision to adapt to a huge range of different environments and lifestyles, and evolve complex behaviors. Much of our knowledge of eye development has been learned from Drosophila, while visual adaptations and behaviors are often more striking and better understood from studies of other insects. However, recent studies in Drosophila and other insects, including bees, beetles, and butterflies, have begun to address this gap by revealing the genetic and developmental bases of differences in eye morphology and key new aspects of compound eye structure and function. Furthermore, technical advances have facilitated the generation of high-resolution connectomic data from different insect species that enhances our understanding of visual information processing, and the impact of changes in these processes on the evolution of vision and behavior. Here, we review these recent breakthroughs and propose that future integrated research from the development to function of visual systems within and among insect species represents a great opportunity to understand the remarkable diversification of insect eyes and vision.


Assuntos
Evolução Biológica , Insetos , Visão Ocular , Animais , Visão Ocular/fisiologia , Insetos/fisiologia , Insetos/genética , Olho/anatomia & histologia , Olho Composto de Artrópodes/fisiologia , Olho Composto de Artrópodes/anatomia & histologia
3.
BMC Biol ; 22(1): 67, 2024 Mar 19.
Artigo em Inglês | MEDLINE | ID: mdl-38504308

RESUMO

BACKGROUND: Insects have evolved complex visual systems and display an astonishing range of adaptations for diverse ecological niches. Species of Drosophila melanogaster subgroup exhibit extensive intra- and interspecific differences in compound eye size. These differences provide an excellent opportunity to better understand variation in insect eye structure and the impact on vision. Here we further explored the difference in eye size between D. mauritiana and its sibling species D. simulans. RESULTS: We confirmed that D. mauritiana have rapidly evolved larger eyes as a result of more and wider ommatidia than D. simulans since they recently diverged approximately 240,000 years ago. The functional impact of eye size, and specifically ommatidia size, is often only estimated based on the rigid surface morphology of the compound eye. Therefore, we used 3D synchrotron radiation tomography to measure optical parameters in 3D, predict optical capacity, and compare the modelled vision to in vivo optomotor responses. Our optical models predicted higher contrast sensitivity for D. mauritiana, which we verified by presenting sinusoidal gratings to tethered flies in a flight arena. Similarly, we confirmed the higher spatial acuity predicted for Drosophila simulans with smaller ommatidia and found evidence for higher temporal resolution. CONCLUSIONS: Our study demonstrates that even subtle differences in ommatidia size between closely related Drosophila species can impact the vision of these insects. Therefore, further comparative studies of intra- and interspecific variation in eye morphology and the consequences for vision among other Drosophila species, other dipterans and other insects are needed to better understand compound eye structure-function and how the diversification of eye size, shape, and function has helped insects to adapt to the vast range of ecological niches.


Assuntos
Drosophila melanogaster , Drosophila , Animais , Drosophila/fisiologia , Drosophila melanogaster/genética , Olho/anatomia & histologia , Especificidade da Espécie
4.
Proc Natl Acad Sci U S A ; 116(38): 19025-19030, 2019 09 17.
Artigo em Inglês | MEDLINE | ID: mdl-31484761

RESUMO

Male genital structures are among the most rapidly evolving morphological traits and are often the only features that can distinguish closely related species. This process is thought to be driven by sexual selection and may reinforce species separation. However, while the genetic bases of many phenotypic differences have been identified, we still lack knowledge about the genes underlying evolutionary differences in male genital organs and organ size more generally. The claspers (surstyli) are periphallic structures that play an important role in copulation in insects. Here, we show that divergence in clasper size and bristle number between Drosophila mauritiana and Drosophila simulans is caused by evolutionary changes in tartan (trn), which encodes a transmembrane leucine-rich repeat domain protein that mediates cell-cell interactions and affinity. There are no fixed amino acid differences in trn between D. mauritiana and D. simulans, but differences in the expression of this gene in developing genitalia suggest that cis-regulatory changes in trn underlie the evolution of clasper morphology in these species. Finally, analyses of reciprocal hemizygotes that are genetically identical, except for the species from which the functional allele of trn originates, determined that the trn allele of D. mauritiana specifies larger claspers with more bristles than the allele of D. simulans Therefore, we have identified a gene underlying evolutionary change in the size of a male genital organ, which will help to better understand not only the rapid diversification of these structures, but also the regulation and evolution of organ size more broadly.


Assuntos
Evolução Biológica , Proteínas de Drosophila/genética , Drosophila melanogaster/anatomia & histologia , Drosophila melanogaster/crescimento & desenvolvimento , Genitália Masculina/anatomia & histologia , Genitália Masculina/crescimento & desenvolvimento , Proteínas de Membrana/genética , Animais , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/genética , Feminino , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Genitália Masculina/metabolismo , Masculino , Proteínas de Membrana/metabolismo , Tamanho do Órgão , Fenótipo
5.
Plant Physiol ; 177(1): 241-254, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29555788

RESUMO

Protein storage vacuoles (PSV) are the main repository of protein in dicotyledonous seeds, but little is known about the origins of these transient organelles. PSV are hypothesized to either arise de novo or originate from the preexisting embryonic vacuole (EV) during seed maturation. Here, we tested these hypotheses by studying PSV formation in Arabidopsis (Arabidopsis thaliana) embryos at different stages of seed maturation and recapitulated this process in Arabidopsis leaves reprogrammed to an embryogenic fate by inducing expression of the LEAFY COTYLEDON2 transcription factor. Confocal and immunoelectron microscopy indicated that both storage proteins and tonoplast proteins typical of PSV were delivered to the preexisting EV in embryos or to the lytic vacuole in reprogrammed leaf cells. In addition, sectioning through embryos at several developmental stages using serial block face scanning electron microscopy revealed the 3D architecture of forming PSV. Our results indicate that the preexisting EV is reprogrammed to become a PSV in Arabidopsis.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/citologia , Proteínas de Armazenamento de Sementes/metabolismo , Sementes/citologia , Vacúolos/metabolismo , Aquaporinas/genética , Aquaporinas/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Corantes Fluorescentes/química , Corantes Fluorescentes/metabolismo , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Concentração de Íons de Hidrogênio , Folhas de Planta/citologia , Folhas de Planta/metabolismo , Plantas Geneticamente Modificadas , Proteínas de Armazenamento de Sementes/genética , Sementes/crescimento & desenvolvimento , Sementes/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
6.
Proc Natl Acad Sci U S A ; 113(2): 452-7, 2016 Jan 12.
Artigo em Inglês | MEDLINE | ID: mdl-26715743

RESUMO

The cytoskeleton is an early attribute of cellular life, and its main components are composed of conserved proteins. The actin cytoskeleton has a direct impact on the control of cell size in animal cells, but its mechanistic contribution to cellular growth in plants remains largely elusive. Here, we reveal a role of actin in regulating cell size in plants. The actin cytoskeleton shows proximity to vacuoles, and the phytohormone auxin not only controls the organization of actin filaments but also impacts vacuolar morphogenesis in an actin-dependent manner. Pharmacological and genetic interference with the actin-myosin system abolishes the effect of auxin on vacuoles and thus disrupts its negative influence on cellular growth. SEM-based 3D nanometer-resolution imaging of the vacuoles revealed that auxin controls the constriction and luminal size of the vacuole. We show that this actin-dependent mechanism controls the relative vacuolar occupancy of the cell, thus suggesting an unanticipated mechanism for cytosol homeostasis during cellular growth.


Assuntos
Actinas/metabolismo , Arabidopsis/citologia , Arabidopsis/crescimento & desenvolvimento , Ácidos Indolacéticos/farmacologia , Vacúolos/metabolismo , Citoesqueleto de Actina/efeitos dos fármacos , Citoesqueleto de Actina/metabolismo , Arabidopsis/efeitos dos fármacos , Imageamento Tridimensional , Membranas Intracelulares/efeitos dos fármacos , Membranas Intracelulares/metabolismo , Meristema/efeitos dos fármacos , Microtúbulos/efeitos dos fármacos , Microtúbulos/metabolismo , Modelos Moleculares , Mutação/genética , Miosinas/metabolismo , Fosfatidilinositóis/metabolismo , Raízes de Plantas/citologia , Raízes de Plantas/efeitos dos fármacos , Estabilidade Proteica/efeitos dos fármacos , Vacúolos/efeitos dos fármacos , Vacúolos/ultraestrutura
7.
Proc Natl Acad Sci U S A ; 110(32): E3007-16, 2013 Aug 06.
Artigo em Inglês | MEDLINE | ID: mdl-23878262

RESUMO

Local recycling of synaptic vesicles (SVs) allows neurons to sustain transmitter release. Extreme activity (e.g., during seizure) may exhaust synaptic transmission and, in vitro, induces bulk endocytosis to recover SV membrane and proteins; how this occurs in animals is unknown. Following optogenetic hyperstimulation of Caenorhabditis elegans motoneurons, we analyzed synaptic recovery by time-resolved behavioral, electrophysiological, and ultrastructural assays. Recovery of docked SVs and of evoked-release amplitudes (indicating readily-releasable pool refilling) occurred within ∼8-20 s (τ = 9.2 s and τ = 11.9 s), whereas locomotion recovered only after ∼60 s (τ = 20 s). During ∼11-s stimulation, 50- to 200-nm noncoated vesicles ("100nm vesicles") formed, which disappeared ∼8 s poststimulation, likely representing endocytic intermediates from which SVs may regenerate. In endophilin, synaptojanin, and dynamin mutants, affecting endocytosis and vesicle scission, resolving 100nm vesicles was delayed (>20 s). In dynamin mutants, 100nm vesicles were abundant and persistent, sometimes continuous with the plasma membrane; incomplete budding of smaller vesicles from 100nm vesicles further implicates dynamin in regenerating SVs from bulk-endocytosed vesicles. Synaptic recovery after exhaustive activity is slow, and different time scales of recovery at ultrastructural, physiological, and behavioral levels indicate multiple contributing processes. Similar processes may jointly account for slow recovery from acute seizures also in higher animals.


Assuntos
Neurônios Motores/fisiologia , Optogenética/métodos , Transmissão Sináptica/fisiologia , Vesículas Sinápticas/fisiologia , Animais , Animais Geneticamente Modificados , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/fisiologia , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/fisiologia , Dinaminas/genética , Dinaminas/metabolismo , Dinaminas/fisiologia , Endocitose/genética , Endocitose/fisiologia , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Microscopia Eletrônica , Microscopia de Fluorescência , Neurônios Motores/metabolismo , Mutação , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Proteínas do Tecido Nervoso/fisiologia , Monoéster Fosfórico Hidrolases/genética , Monoéster Fosfórico Hidrolases/metabolismo , Monoéster Fosfórico Hidrolases/fisiologia , Interferência de RNA , Vesículas Sinápticas/metabolismo , Vesículas Sinápticas/ultraestrutura , Fatores de Tempo
8.
Proc Natl Acad Sci U S A ; 110(13): 4986-91, 2013 Mar 26.
Artigo em Inglês | MEDLINE | ID: mdl-23457265

RESUMO

Mutations in the Tar DNA binding protein of 43 kDa (TDP-43; TARDBP) are associated with amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration with TDP-43(+) inclusions (FTLD-TDP). To determine the physiological function of TDP-43, we knocked out zebrafish Tardbp and its paralogue Tardbp (TAR DNA binding protein-like), which lacks the glycine-rich domain where ALS- and FTLD-TDP-associated mutations cluster. tardbp mutants show no phenotype, a result of compensation by a unique splice variant of tardbpl that additionally contains a C-terminal elongation highly homologous to the glycine-rich domain of tardbp. Double-homozygous mutants of tardbp and tardbpl show muscle degeneration, strongly reduced blood circulation, mispatterning of vessels, impaired spinal motor neuron axon outgrowth, and early death. In double mutants the muscle-specific actin binding protein Filamin Ca is up-regulated. Strikingly, Filamin C is similarly increased in the frontal cortex of FTLD-TDP patients, suggesting aberrant expression in smooth muscle cells and TDP-43 loss-of-function as one underlying disease mechanism.


Assuntos
Axônios/metabolismo , Proteínas de Ligação a DNA , Neurônios Motores/metabolismo , Atrofia Muscular/metabolismo , Mutação , Doenças Vasculares/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/metabolismo , Esclerose Lateral Amiotrófica/genética , Esclerose Lateral Amiotrófica/metabolismo , Esclerose Lateral Amiotrófica/patologia , Animais , Axônios/patologia , Proteínas Contráteis/genética , Proteínas Contráteis/metabolismo , Filaminas , Humanos , Proteínas dos Microfilamentos/genética , Proteínas dos Microfilamentos/metabolismo , Neurônios Motores/patologia , Atrofia Muscular/genética , Atrofia Muscular/patologia , Estrutura Terciária de Proteína , Doenças Vasculares/genética , Doenças Vasculares/patologia , Proteínas de Peixe-Zebra/genética
9.
Proc Natl Acad Sci U S A ; 109(15): 5862-7, 2012 Apr 10.
Artigo em Inglês | MEDLINE | ID: mdl-22451907

RESUMO

Presynaptic nerve terminals are formed from preassembled vesicles that are delivered to the prospective synapse by kinesin-mediated axonal transport. However, precisely how the various cargoes are linked to the motor proteins remains unclear. Here, we report a transport complex linking syntaxin 1a (Stx) and Munc18, two proteins functioning in synaptic vesicle exocytosis at the presynaptic plasma membrane, to the motor protein Kinesin-1 via the kinesin adaptor FEZ1. Mutation of the FEZ1 ortholog UNC-76 in Caenorhabditis elegans causes defects in the axonal transport of Stx. We also show that binding of FEZ1 to Kinesin-1 and Munc18 is regulated by phosphorylation, with a conserved site (serine 58) being essential for binding. When expressed in C. elegans, wild-type but not phosphorylation-deficient FEZ1 (S58A) restored axonal transport of Stx. We conclude that FEZ1 operates as a kinesin adaptor for the transport of Stx, with cargo loading and unloading being regulated by protein kinases.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Transporte Axonal , Proteínas de Caenorhabditis elegans/metabolismo , Cinesinas/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Neuropeptídeos/metabolismo , Sintaxina 1/metabolismo , Animais , Axônios/metabolismo , Caenorhabditis elegans/metabolismo , Células HEK293 , Humanos , Proteínas Munc18/metabolismo , Proteínas Mutantes/metabolismo , Mutação/genética , Fosforilação , Ligação Proteica , Transporte Proteico
10.
Methods Mol Biol ; 2772: 15-25, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38411803

RESUMO

The endoplasmic reticulum (ER) forms an extensive network in plant cells. In leaf cells and vacuolated root cells it is mainly restricted to the cortex, whereas in the root meristem the cortical and cytoplasmic ER takes up a large volume throughout the entire cell. Only 3D electron microscopy provides sufficient resolution to understand the spatial organization of the ER in the root. Here we present two protocols for 3D EM imaging of the ER across a range of scales. For large-scale ER structure analysis, we describe selective ER staining with ZIO that allows for automated or semi-automated ER segmentation. For smaller regions of ER, we describe high-pressure freezing, which enables almost instantaneous fixation of plant tissues but without organelle specific staining. These fixation and staining techniques are suitable for a range of imaging modalities, including serial sections, array tomography, serial block face-scanning electron microscopy (SBF-SEM), or focused ion beam (FIB) SEM.


Assuntos
Elétrons , Retículo Endoplasmático , Microscopia Eletrônica , Citosol , Técnicas Histológicas
11.
Cell Rep ; 43(3): 113791, 2024 Mar 26.
Artigo em Inglês | MEDLINE | ID: mdl-38428420

RESUMO

The "ribbon," a structural arrangement in which Golgi stacks connect to each other, is considered to be restricted to vertebrate cells. Although ribbon disruption is linked to various human pathologies, its functional role in cellular processes remains unclear. In this study, we investigate the evolutionary origin of the Golgi ribbon. We observe a ribbon-like architecture in the cells of several metazoan taxa suggesting its early emergence in animal evolution predating the appearance of vertebrates. Supported by AlphaFold2 modeling, we propose that the evolution of Golgi reassembly and stacking protein (GRASP) binding by golgin tethers may have driven the joining of Golgi stacks resulting in the ribbon-like configuration. Additionally, we find that Golgi ribbon assembly is a shared developmental feature of deuterostomes, implying a role in embryogenesis. Overall, our study points to the functional significance of the Golgi ribbon beyond vertebrates and underscores the need for further investigations to unravel its elusive biological roles.


Assuntos
Complexo de Golgi , Proteínas de Membrana , Animais , Humanos , Proteínas de Membrana/metabolismo , Complexo de Golgi/metabolismo , Citoesqueleto/metabolismo , Células HeLa , Vertebrados
12.
Science ; 380(6642): 293-297, 2023 04 21.
Artigo em Inglês | MEDLINE | ID: mdl-37079688

RESUMO

A fundamental breakthrough in neurobiology has been the formulation of the neuron doctrine by Santiago Ramón y Cajal, which stated that the nervous system is composed of discrete cells. Electron microscopy later confirmed the doctrine and allowed the identification of synaptic connections. In this work, we used volume electron microscopy and three-dimensional reconstructions to characterize the nerve net of a ctenophore, a marine invertebrate that belongs to one of the earliest-branching animal lineages. We found that neurons in the subepithelial nerve net have a continuous plasma membrane that forms a syncytium. Our findings suggest fundamental differences of nerve net architectures between ctenophores and cnidarians or bilaterians and offer an alternative perspective on neural network organization and neurotransmission.


Assuntos
Evolução Biológica , Ctenóforos , Sistema Nervoso , Animais , Rede Nervosa/fisiologia , Neurônios/fisiologia , Transmissão Sináptica
13.
Sci Rep ; 12(1): 6402, 2022 04 17.
Artigo em Inglês | MEDLINE | ID: mdl-35431314

RESUMO

The coexistence of different mating strategies, whereby a species can reproduce both by selfing and outcrossing, is an evolutionary enigma. Theory predicts two predominant stable mating states: outcrossing with strong inbreeding depression or selfing with weak inbreeding depression. As these two mating strategies are subject to opposing selective forces, mixed breeding systems are thought to be a rare transitory state yet can persist even after multiple speciation events. We hypothesise that if each mating strategy plays a distinctive role during some part of the species life history, opposing selective pressures could be balanced, permitting the stable co-existence of selfing and outcrossing sexual morphs. In this scenario, we would expect each morph to be specialised in their respective roles. Here we show, using behavioural, physiological and gene expression studies, that the selfing (hermaphrodite) and outcrossing (female) sexual morphs of the trioecious nematode Auanema freiburgensis have distinct adaptations optimised for their different roles during the life cycle. A. freiburgensis hermaphrodites are known to be produced under stressful conditions and are specialised for dispersal to new habitat patches. Here we show that they exhibit metabolic and intestinal changes enabling them to meet the cost of dispersal and reproduction. In contrast, A. freiburgensis females are produced in favourable conditions and facilitate rapid population growth. We found that females compensate for the lack of reproductive assurance by reallocating resources from intestinal development to mate-finding behaviour. The specialisation of each mating system for its role in the life cycle could balance opposing selective forces allowing the stable maintenance of both mating systems in A. freiburgensis.


Assuntos
Evolução Biológica , Depressão por Endogamia , Animais , Feminino , Estágios do Ciclo de Vida , Reprodução/fisiologia
14.
Curr Biol ; 31(23): 5274-5285.e6, 2021 12 06.
Artigo em Inglês | MEDLINE | ID: mdl-34587474

RESUMO

Ctenophores are gelatinous marine animals famous for locomotion by ciliary combs. Due to the uncertainties of the phylogenetic placement of ctenophores and the absence of some key bilaterian neuronal genes, it has been hypothesized that their neurons evolved independently. Additionally, recent whole-body, single-cell RNA sequencing (scRNA-seq) analysis failed to identify ctenophore neurons using any of the known neuronal molecular markers. To reveal the molecular machinery of ctenophore neurons, we have characterized the neuropeptide repertoire of the ctenophore Mnemiopsis leidyi. Using the machine learning NeuroPID tool, we predicted 129 new putative neuropeptide precursors. Sixteen of them were localized to the subepithelial nerve net (SNN), sensory aboral organ (AO), and epithelial sensory cells (ESCs), providing evidence that they are neuropeptide precursors. Four of these putative neuropeptides had a behavioral effect and increased the animals' swimming speed. Intriguingly, these putative neuropeptides finally allowed us to identify neuronal cell types in single-cell transcriptomic data and reveal the molecular identity of ctenophore neurons. High-resolution electron microscopy and 3D reconstructions of the nerve net underlying the comb plates confirmed a more than 100-year-old hypothesis of anastomoses between neurites of the same cell in ctenophores and revealed that they occur through a continuous membrane. Our work demonstrates the unique ultrastructure of the peptidergic nerve net and a rich neuropeptide repertoire of ctenophores, supporting the hypothesis that the first nervous system(s) evolved as nets of peptidergic cells.


Assuntos
Ctenóforos , Neuropeptídeos , Animais , Ctenóforos/anatomia & histologia , Sistema Nervoso/metabolismo , Neurônios , Neuropeptídeos/genética , Neuropeptídeos/metabolismo , Filogenia
15.
G3 (Bethesda) ; 10(3): 1005-1018, 2020 03 05.
Artigo em Inglês | MEDLINE | ID: mdl-31919111

RESUMO

The compound eyes of insects exhibit striking variation in size, reflecting adaptation to different lifestyles and habitats. However, the genetic and developmental bases of variation in insect eye size is poorly understood, which limits our understanding of how these important morphological differences evolve. To address this, we further explored natural variation in eye size within and between four species of the Drosophila melanogaster species subgroup. We found extensive variation in eye size among these species, and flies with larger eyes generally had a shorter inter-ocular distance and vice versa We then carried out quantitative trait loci (QTL) mapping of intra-specific variation in eye size and inter-ocular distance in both D. melanogaster and D. simulans This revealed that different genomic regions underlie variation in eye size and inter-ocular distance in both species, which we corroborated by introgression mapping in D. simulans This suggests that although there is a trade-off between eye size and inter-ocular distance, variation in these two traits is likely to be caused by different genes and so can be genetically decoupled. Finally, although we detected QTL for intra-specific variation in eye size at similar positions in D. melanogaster and D. simulans, we observed differences in eye fate commitment between strains of these two species. This indicates that different developmental mechanisms and therefore, most likely, different genes contribute to eye size variation in these species. Taken together with the results of previous studies, our findings suggest that the gene regulatory network that specifies eye size has evolved at multiple genetic nodes to give rise to natural variation in this trait within and among species.


Assuntos
Drosophila melanogaster/anatomia & histologia , Drosophila melanogaster/genética , Drosophila simulans/anatomia & histologia , Drosophila simulans/genética , Olho/anatomia & histologia , Animais , Feminino , Genótipo , Masculino , Tamanho do Órgão/genética , Fenótipo , Locos de Características Quantitativas
16.
Dev Genes Evol ; 219(8): 399-407, 2009 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-19760181

RESUMO

The genetic control of leg development is well characterized in the fly Drosophila melanogaster. These control mechanisms, however, must differ to some degree between different insect species to account for the morphological diversity of thoracic legs in the insects. The legs of the flour beetle Tribolium castaneum differ from the Drosophila legs in their developmental mode as well as in their specific morphology especially at the larval stage. In order to identify genes involved in the morphogenesis of the Tribolium larval legs, we have analyzed EGFP enhancer trap lines of Tribolium. We have identified the zfh2 gene as a novel factor required for normal leg development in Tribolium. RNA interference with zfh2 function leads to two alternative classes of leg phenotype. The loss of a leg segment boundary and the generation of ectopic outgrowths in one class of phenotype suggest a role in leg segmentation and segment growth. The malformation of the pretarsal claw in the second class of phenotype suggests a role in distal development and the morphogenesis of the claw-shaped morphology of the pretarsus. This suggests that zfh2 is involved in the regulation of an unidentified target gene in a concentration-dependent manner. Our results demonstrate that enhancer trap screens in T. castaneum have the potential to identify novel gene functions regulating specific developmental processes.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Genes de Insetos , Tribolium/crescimento & desenvolvimento , Tribolium/genética , Animais , Padronização Corporal , Proteínas de Ligação a DNA/genética , Extremidades/crescimento & desenvolvimento , Regulação da Expressão Gênica no Desenvolvimento , Proteínas de Fluorescência Verde/metabolismo , Larva/crescimento & desenvolvimento , Mutagênese Insercional
17.
Nat Commun ; 10(1): 984, 2019 02 28.
Artigo em Inglês | MEDLINE | ID: mdl-30816109

RESUMO

The endoplasmic reticulum (ER) is a highly dynamic polygonal membrane network composed of interconnected tubules and sheets (cisternae) that forms the first compartment in the secretory pathway involved in protein translocation, folding, glycosylation, quality control, lipid synthesis, calcium signalling, and metabolon formation. Despite its central role in this plethora of biosynthetic, metabolic and physiological processes, there is little quantitative information on ER structure, morphology or dynamics. Here we describe a software package (AnalyzER) to automatically extract ER tubules and cisternae from multi-dimensional fluorescence images of plant ER. The structure, topology, protein-localisation patterns, and dynamics are automatically quantified using spatial, intensity and graph-theoretic metrics. We validate the method against manually-traced ground-truth networks, and calibrate the sub-resolution width estimates against ER profiles identified in serial block-face SEM images. We apply the approach to quantify the effects on ER morphology of drug treatments, abiotic stress and over-expression of ER tubule-shaping and cisternal-modifying proteins.


Assuntos
Retículo Endoplasmático/ultraestrutura , Plantas/ultraestrutura , Software , Arabidopsis/genética , Arabidopsis/metabolismo , Arabidopsis/ultraestrutura , Retículo Endoplasmático/genética , Retículo Endoplasmático/metabolismo , Processamento de Imagem Assistida por Computador , Microscopia Eletrônica de Varredura , Microscopia de Fluorescência , Proteínas de Plantas/metabolismo , Plantas/genética , Plantas/metabolismo , Plantas Geneticamente Modificadas , Nicotiana/genética , Nicotiana/metabolismo , Nicotiana/ultraestrutura
18.
Methods Mol Biol ; 1691: 15-21, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29043666

RESUMO

The endoplasmic reticulum (ER) forms an extensive network in plant cells. In leaf cells and vacuolated root cells it is mainly restricted to the cortex whereas in the root meristem the cortical and cytoplasmic ER takes up a large volume throughout the entire cell. Only 3D electron microscopy provides sufficient resolution to understand the spatial organization of the ER in the root. However, high contrast staining and optimally ER specific staining is essential. Here we describe a protocol for selective ER staining that allows automated or semiautomated segmentation of the organelle in 3D datasets obtained from serial sections, Array Tomography, Serial Block Face Scanning Electron Microscopy (SBFSEM), or Focused Ion Beam (FIB) SEM.


Assuntos
Retículo Endoplasmático/ultraestrutura , Imageamento Tridimensional , Microscopia Eletrônica , Imageamento Tridimensional/métodos , Microscopia Eletrônica/métodos , Células Vegetais
19.
Sci Rep ; 8(1): 2310, 2018 02 02.
Artigo em Inglês | MEDLINE | ID: mdl-29396477

RESUMO

Reticulons are integral ER membrane proteins characterised by a reticulon homology domain comprising four transmembrane domains which results in the proteins sitting in the membrane in a W-topology. Here we report on a novel subgroup of reticulons with an extended N-terminal domain and in particular on arabidopsis reticulon 20. Using high resolution confocal microscopy we show that reticulon 20 is located in a unique punctate pattern on the ER membrane. Its closest homologue reticulon 19 labels the whole ER. Other than demonstrated for the other members of the reticulon protein family RTN20 and 19 do not display ER constriction phenotypes on over expression. We show that mutants in RTN20 or RTN19, respectively, display a significant change in sterol composition in roots indicating a role in lipid regulation. A third homologue in this family -3BETAHSD/D1- is unexpectedly localised to ER exit sites resulting in an intriguing location difference for the three proteins.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Retículo Endoplasmático/metabolismo , Metabolismo dos Lipídeos , Proteínas de Membrana/metabolismo , Microscopia Confocal , Microscopia de Fluorescência
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