RESUMEN
Autophagy is a conserved intracellular degradation pathway that generates de novo double-membrane autophagosomes to target a wide range of material for lysosomal degradation. In multicellular organisms, autophagy initiation requires the timely assembly of a contact site between the ER and the nascent autophagosome. Here, we report the in vitro reconstitution of a full-length seven-subunit human autophagy initiation supercomplex built on a core complex of ATG13-101 and ATG9. Assembly of this core complex requires the rare ability of ATG13 and ATG101 to switch between distinct folds. The slow spontaneous metamorphic conversion is rate limiting for the self-assembly of the supercomplex. The interaction of the core complex with ATG2-WIPI4 enhances tethering of membrane vesicles and accelerates lipid transfer of ATG2 by both ATG9 and ATG13-101. Our work uncovers the molecular basis of the contact site and its assembly mechanisms imposed by the metamorphosis of ATG13-101 to regulate autophagosome biogenesis in space and time.
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Autofagosomas , Autofagia , Humanos , Proteínas Relacionadas con la Autofagia/genética , Proteínas Relacionadas con la Autofagia/metabolismo , Autofagia/fisiología , Autofagosomas/metabolismo , Proteínas de la Membrana/metabolismo , LípidosRESUMEN
In the body of multicellular organisms, macrophages play an indispensable role in maintaining tissue homeostasis by removing old, apoptotic and damaged cells. In addition, macrophages allow significant remodeling of body plans during embryonic morphogenesis, regeneration and metamorphosis. Although the huge amount of organic matter that must be removed during these processes represents a potential source of nutrients, their further use by the organism has not yet been addressed. Here, we document that, during metamorphosis, Drosophila larval adipose tissue is infiltrated by macrophages, which remove dying adipocytes by efferocytosis and engulf leaking RNA-protein granules and lipids. Consequently, the infiltrating macrophages transiently adopt the adipocyte-like metabolic profile to convert remnants of dying adipocytes to lipoproteins and storage peptides that nutritionally support post-metamorphic development. This process is fundamental for the full maturation of ovaries and the achievement of early fecundity of individuals. Whether macrophages play an analogous role in other situations of apoptotic cell removal remains to be elucidated.
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Drosophila , Macrófagos , Humanos , Animales , Macrófagos/metabolismo , Tejido Adiposo/metabolismo , Adipocitos/metabolismoRESUMEN
Autophagy is essential for the turnover of damaged organelles and long-lived proteins. It is responsible for many biological processes such as maintaining brain functions and aging. Impaired autophagy is often linked to neurodevelopmental and neurodegenerative diseases in humans. However, the role of autophagy in neuronal pruning during development remains poorly understood. Here, we report that autophagy regulates dendrite-specific pruning of ddaC sensory neurons in parallel to local caspase activation. Impaired autophagy causes the formation of ubiquitinated protein aggregates in ddaC neurons, dependent on the autophagic receptor Ref(2)P. Furthermore, the metabolic regulator AMP-activated protein kinase and the insulin-target of rapamycin pathway act upstream to regulate autophagy during dendrite pruning. Importantly, autophagy is required to activate the transcription factor CncC (Cap "n" collar isoform C), thereby promoting dendrite pruning. Conversely, CncC also indirectly affects autophagic activity via proteasomal degradation, as impaired CncC results in the inhibition of autophagy through sequestration of Atg8a into ubiquitinated protein aggregates. Thus, this study demonstrates the important role of autophagy in activating CncC prior to dendrite pruning, and further reveals an interplay between autophagy and CncC in neuronal pruning.
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Proteínas de Drosophila , Drosophila , Compuestos de Amonio Cuaternario , Animales , Humanos , Autofagia/fisiología , Dendritas/metabolismo , Drosophila/metabolismo , Proteínas de Drosophila/metabolismo , Plasticidad Neuronal , Proteínas Ubiquitinadas/metabolismoRESUMEN
The prothoracic gland (PG) is a major insect endocrine organ. It is the principal source of insect steroid hormones, and critical for key developmental events such as the molts, the establishment of critical weight (CW), pupation, and sexual maturation. However, little is known about the developmental processes that regulate PG morphology. In this study, we identified soul, which encodes a PG-specific basic helix-loop-helix (bHLH) transcription factor. We demonstrate that Tap, also a bHLH protein, dimerizes with Soul. Both are expressed in the developing PG. Interfering with either soul or tap function caused strikingly similar phenotypes, resulting in small and fragmented PGs, the abolishment of steroid hormone-producing gene expression, larval arrest, and a failure to undergo metamorphosis. Furthermore, both soul and tap showed expression peaks just prior to the CW checkpoint. Disrupting soul- or tap-function before, but not after, the CW checkpoint caused larval arrest, and perturbed highly similar gene cohorts, which were enriched for regulators and components of the steroid hormone biosynthesis pathway. Intriguingly, a chitin-based cuticle gene, Cpr49Ah, and a POU domain transcription factor gene, pdm3, are direct target genes of the Soul/Tap complex, and disruption of either phenocopied key aspects of soul/tap loss-of-function phenotypes. Taken together, our findings demonstrate that the Soul/Tap heterodimer resides at the top of a complex gene hierarchy that drives PG development, CW establishment, and steroid hormone production.
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Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico , Proteínas de Drosophila , Drosophila melanogaster , Regulación del Desarrollo de la Expresión Génica , Larva , Animales , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Larva/genética , Larva/crecimiento & desarrollo , Larva/metabolismo , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Drosophila melanogaster/crecimiento & desarrollo , Glándulas Endocrinas/metabolismo , Metamorfosis Biológica/genéticaRESUMEN
Many insects are dependent on microbial mutualists, which are often harbored in specialized symbiotic organs. Upon metamorphosis, insect organs are drastically reorganized. What mechanism regulates the remodeling of the symbiotic organ upon metamorphosis? How does it affect the microbial symbiont therein? Here, we addressed these fundamental issues of symbiosis by experimentally manipulating insect metamorphosis. The stinkbug Plautia stali possesses a midgut symbiotic organ wherein an essential bacterial symbiont resides. By RNAi of master regulator genes for metamorphosis, Kr-h1 over nymphal traits and E93 over adult traits, we generated precocious adults and supernumerary nymphs of P. stali, thereby disentangling the effects of metamorphosis, growth level, developmental stage, and other factors on the symbiotic system. Upon metamorphosis, the symbiotic organ of P. stali was transformed from nymph type to adult type. The supernumerary nymphs and the precocious adults, respectively, developed nymph-type and adult-type symbiotic organs not only morphologically but also transcriptomically, uncovering that metamorphic remodeling of the symbiotic organ is under the control of the MEKRE93 pathway. Transcriptomic, cytological, and biochemical analyses unveiled that the structural and transcriptomic remodeling of the symbiotic organ toward adult emergence underpins its functional extension to food digestion in addition to the original role of symbiont retention for essential nutrient production. Notably, we found that the symbiotic bacteria in the adult-type symbiotic organ up-regulated genes for production of sulfur-containing essential amino acids, methionine and cysteine, that are rich in eggs and sperm, uncovering adult-specific symbiont functioning for host reproduction and highlighting intricate host-symbiont interactions associated with insect metamorphosis.
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Heterópteros , Simbiosis , Masculino , Animales , Simbiosis/fisiología , Semen , Sistema Digestivo/microbiología , Insectos , Heterópteros/fisiología , Bacterias/genética , Metamorfosis BiológicaRESUMEN
The early embryo of the cockroach Blattella germanica exhibits high E93 expression. In general, E93 triggers adult morphogenesis during postembryonic development. Here we show that E93 is also crucial in early embryogenesis in the cockroach, as a significant number of E93-depleted embryos are unable to develop the germ band under maternal RNAi treatment targeting E93. Moreover, transcriptomic analysis indicates that E93 depletion results in important gene expression changes in the early embryo, and many of the differentially expressed genes are involved in development. Then, using public databases, we gathered E93 expression data in embryo and preadult stages, finding that embryonic expression of E93 is high in hemimetabolan species (whose juveniles, or nymphs, are similar to the adult) and low in holometabolans (whose juveniles, or larvae, are different from the adult). E93 expression is also low in Thysanoptera and in Hemiptera Sternorrhyncha, hemimetabolans with postembryonic quiescent stages, as well as in Odonata, the nymph of which is very different from the adult. In ametabolans, such as the Zygentoma Thermobia domestica, E93 transcript levels are very high in the early embryo, whereas during postembryonic development they are medium and relatively constant. We propose the hypothesis that during evolution, a reduction of E93 expression in the embryo of hemimetabolans facilitated the larval development and the emergence of holometaboly. Independent decreases of E93 transcripts in the embryo of Odonata, Thysanoptera, and different groups of Hemiptera Sternorrhyncha would have allowed the development of modified juvenile stages adapted to specific ecophysiological conditions.
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Hemípteros , Insectos , Animales , Insectos/metabolismo , Metamorfosis Biológica/genética , Larva , Hemípteros/genética , Interferencia de ARN , Regulación del Desarrollo de la Expresión Génica , Proteínas de Insectos/genéticaRESUMEN
Apoptosis is a regulated cell death that depends on caspases. It has mainly been studied as a mechanism for the removal of unwanted cells. However, apoptotic cells can induce fate or behavior changes of their neighbors and thereby participate in development. Here, we address the functions of apoptosis during metamorphosis of the cnidarian Hydractinia symbiolongicarpus. We describe the apoptotic profile during metamorphosis of the larva and identify Caspase3/7a, but no other executioner caspases, as essential for apoptosis in this context. Using pharmacological and genetic approaches, we find that apoptosis is required for normal head development. Inhibition of apoptosis resulted in defects in head morphogenesis. Neurogenesis was compromised in the body column of apoptosis-inhibited animals but there was no effect on the survival or proliferation of stem cells, suggesting that apoptosis is required for cellular commitment rather than for the maintenance of their progenitors. Differential transcriptomic analysis identifies TRAF genes as downregulated in apoptosis-inhibited larvae and functional experiments provide evidence that they are essential for head development. Finally, we find no major role for apoptosis in head regeneration in this animal, in contrast to the significance of apoptosis in Hydra head regeneration.
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Apoptosis , Cabeza , Metamorfosis Biológica , Animales , Apoptosis/genética , Caspasas/metabolismo , Caspasas/genética , Larva/crecimiento & desarrollo , Neurogénesis/genética , Hidrozoos/genética , Hidrozoos/crecimiento & desarrollo , Regulación del Desarrollo de la Expresión Génica , Regeneración/genética , Regeneración/fisiología , Cnidarios/genética , Células Madre/metabolismo , Células Madre/citologíaRESUMEN
The development of the sea urchin larval body plan is well understood from extensive studies of embryonic patterning. However, fewer studies have investigated the late larval stages during which the unique pentaradial adult body plan develops. Previous work on late larval development highlights major tissue changes leading up to metamorphosis, but the location of specific cell types during juvenile development is less understood. Here, we improve on technical limitations by applying highly sensitive hybridization chain reaction fluorescent in situ hybridization (HCR-FISH) to the fast-developing and transparent sea urchin Lytechinus pictus, with a focus on skeletogenic cells. First, we show that HCR-FISH can be used in L. pictus to precisely localize skeletogenic cells in the rudiment. In doing so, we provide a detailed staging scheme for the appearance of skeletogenic cells around the rudiment prior to and during biomineralization and show that many skeletogenic cells unassociated with larval rods localize outside of the rudiment prior to localizing inside. Second, we show that downstream biomineralization genes have similar expression patterns during larval and juvenile skeletogenesis, suggesting some conservation of skeletogenic mechanisms during development between stages. Third, we find co-expression of blastocoelar and skeletogenic cell markers around juvenile skeleton located outside of the rudiment, which is consistent with data showing that cells from the non-skeletogenic mesoderm embryonic lineage contribute to the juvenile skeletogenic cell lineage. This work sets the foundation for subsequent studies of other cell types in the late larva of L. pictus to better understand juvenile body plan development, patterning, and evolution.
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Larva , Lytechinus , Animales , Lytechinus/embriología , Larva/crecimiento & desarrollo , Regulación del Desarrollo de la Expresión Génica , Hibridación Fluorescente in Situ , Erizos de Mar/embriología , Metamorfosis Biológica , Tipificación del Cuerpo/genética , BiomineralizaciónRESUMEN
Many insects undergo the process of metamorphosis when larval precursor cells begin to differentiate to create the adult body. The larval precursor cells retain stem cell-like properties and contribute to the regenerative ability of larval appendages. Here we demonstrate that two Broad-complex/Tramtrack/Bric-à-brac Zinc-finger (BTB) domain transcription factors, Chronologically inappropriate morphogenesis (Chinmo) and Abrupt (Ab), act cooperatively to repress metamorphosis in the flour beetle, Tribolium castaneum. Knockdown of chinmo led to precocious development of pupal legs and antennae. We show that although topical application of juvenile hormone (JH) prevents the decrease in chinmo expression in the final instar, chinmo and JH act in distinct pathways. Another gene encoding the BTB domain transcription factor, Ab, was also necessary for the suppression of broad (br) expression in T. castaneum in a chinmo RNAi background, and simultaneous knockdown of ab and chinmo led to the precocious onset of metamorphosis. Furthermore, knockdown of ab led to the loss of regenerative potential of larval legs independently of br. In contrast, chinmo knockdown larvae exhibited pupal leg regeneration when a larval leg was ablated. Taken together, our results show that both ab and chinmo are necessary for the maintenance of the larval tissue identity and, apart from its role in repressing br, ab acts as a crucial regulator of larval leg regeneration. Our findings indicate that BTB domain proteins interact in a complex manner to regulate larval and pupal tissue homeostasis.
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Escarabajos , Metamorfosis Biológica , Morfogénesis , Factores de Transcripción , Tribolium , Animales , Escarabajos/metabolismo , Regulación del Desarrollo de la Expresión Génica , Proteínas de Insectos/genética , Proteínas de Insectos/metabolismo , Hormonas Juveniles , Larva/metabolismo , Metamorfosis Biológica/genética , Morfogénesis/genética , Pupa/metabolismo , Factores de Transcripción/metabolismo , Tribolium/genética , Regeneración/genéticaRESUMEN
During anuran metamorphosis from herbivorous tadpoles to carnivorous frogs, the gastrointestinal (GI) tract undergoes drastic remodeling, such as the formation of the stomach-intestine boundary and the development of the pyloric sphincter at the posterior end of the stomach. However, the morphogenetic process and molecular mechanisms of how the pyloric sphincter is formed during metamorphosis, instead of during embryogenesis as in amniotes, are largely uninvestigated. Using the African clawed frog Xenopus laevis, we histologically examined the development of the pylorus region from embryonic to froglet stages and performed spatiotemporal gene expression analyses. We found that the pyloric sphincter is formed at a flexure within the pyloric region during metamorphic climax, and that the pyloric and duodenal epithelia, which are morphologically indistinguishable before sphincter formation, become clearly demarcated by the sphincter at the end of metamorphosis. Consistent with these morphological changes, expression domains of a stomach marker barx1 and an intestine marker cdx2 overlapped until late metamorphic climax, but became separated after metamorphosis. Despite the absence of the sphincter before metamorphosis, various genes crucial for sphincter formation in amniotes were already expressed in the pylorus region of Xenopus embryos. RNA-sequencing analysis at pre-metamorphic and metamorphic-climax stages suggest unappreciated roles of genes, such as those for retinoic acid signaling and various transcription factors, in suppressing or promoting sphincter formation. These data provide histological and molecular insights into the heterochrony of the pyloric sphincter formation in amniotes and anurans.
RESUMEN
Proven roles for hemocytes (blood cells) have expanded beyond the control of infections in Drosophila. Despite this, the crucial role of hemocytes in post-embryonic development has long thought to be limited to control of microorganisms during metamorphosis. This has previously been shown by rescue of adult development in hemocyte-ablation models under germ-free conditions. Here, we show that hemocytes have an essential role in post-embryonic development beyond their ability to control the microbiota. Using a newly generated strong hemocyte-specific driver line for the GAL4/UAS system, we show that specific ablation of hemocytes is early pupal lethal, even under axenic conditions. Genetic rescue experiments prove that this is a hemocyte-specific phenomenon. RNA-seq data suggests that dysregulation of the midgut is a prominent consequence of hemocyte ablation in larval stages, resulting in reduced gut lengths. Dissection suggests that multiple processes may be affected during metamorphosis. We believe this previously unreported role for hemocytes during metamorphosis is a major finding for the field.
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Proteínas de Drosophila , Microbiota , Animales , Drosophila , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Desarrollo Embrionario , Hemocitos , LarvaRESUMEN
During Drosophila metamorphosis, the ddaC dendritic arborisation sensory neurons selectively prune their larval dendrites in response to steroid hormone ecdysone signalling. The Nrf2-Keap1 pathway acts downstream of ecdysone signalling to promote proteasomal degradation and thereby dendrite pruning. However, how the Nrf2-Keap1 pathway is activated remains largely unclear. Here, we demonstrate that the metabolic regulator AMP-activated protein kinase (AMPK) plays a cell-autonomous role in dendrite pruning. Importantly, AMPK is required for Mical and Headcase expression and for activation of the Nrf2-Keap1 pathway. We reveal that AMPK promotes the Nrf2-Keap1 pathway and dendrite pruning partly via inhibition of the insulin pathway. Moreover, the AMPK-insulin pathway is required for ecdysone signalling to activate the Nrf2-Keap1 pathway during dendrite pruning. Overall, this study reveals an important mechanism whereby ecdysone signalling activates the Nrf2-Keap1 pathway via the AMPK-insulin pathway to promote dendrite pruning, and further suggests that during the nonfeeding prepupal stage metabolic alterations lead to activation of the Nrf2-Keap1 pathway and dendrite pruning.
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Proteínas de Drosophila , Insulinas , Proteínas Quinasas Activadas por AMP/genética , Proteínas Quinasas Activadas por AMP/metabolismo , Animales , Dendritas/metabolismo , Drosophila/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Ecdisona/metabolismo , Regulación del Desarrollo de la Expresión Génica , Insulinas/metabolismo , Proteína 1 Asociada A ECH Tipo Kelch/genética , Proteína 1 Asociada A ECH Tipo Kelch/metabolismo , Factor 2 Relacionado con NF-E2/genética , Factor 2 Relacionado con NF-E2/metabolismo , Plasticidad NeuronalRESUMEN
The evolutionarily conserved Glycogen Synthase Kinase 3ß (GSK3ß), a negative regulator of microtubules, is crucial for neuronal polarization, growth and migration during animal development. However, it remains unknown whether GSK3ß regulates neuronal pruning, which is a regressive process. Here, we report that the Drosophila GSK3ß homologue Shaggy (Sgg) is cell-autonomously required for dendrite pruning of ddaC sensory neurons during metamorphosis. Sgg is necessary and sufficient to promote microtubule depolymerization, turnover and disassembly in the dendrites. Although Sgg is not required for the minus-end-out microtubule orientation in dendrites, hyperactivated Sgg can disturb the dendritic microtubule orientation. Moreover, our pharmacological and genetic data suggest that Sgg is required to promote dendrite pruning at least partly via microtubule disassembly. We show that Sgg and Par-1 kinases act synergistically to promote microtubule disassembly and dendrite pruning. Thus, Sgg and Par-1 might converge on and phosphorylate a common downstream microtubule-associated protein(s) to disassemble microtubules and thereby facilitate dendrite pruning.
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Proteínas de Drosophila , Drosophila , Animales , Drosophila/genética , Proteínas de Drosophila/genética , Glucógeno Sintasa Quinasa 3 beta/genética , Dendritas/genética , Células Receptoras Sensoriales , Microtúbulos , Plasticidad Neuronal/genética , Drosophila melanogaster/genéticaRESUMEN
Normal tables of development are essential for studies of embryogenesis, serving as an important resource for model organisms, including the frog Xenopus laevis. Xenopus has long been used to study developmental and cell biology, and is an increasingly important model for human birth defects and disease, genomics, proteomics and toxicology. Scientists utilize Nieuwkoop and Faber's classic 'Normal Table of Xenopus laevis (Daudin)' and accompanying illustrations to enable experimental reproducibility and reuse the illustrations in new publications and teaching. However, it is no longer possible to obtain permission for these copyrighted illustrations. We present 133 new, high-quality illustrations of X. laevis development from fertilization to metamorphosis, with additional views that were not available in the original collection. All the images are available on Xenbase, the Xenopus knowledgebase (http://www.xenbase.org/entry/zahn.do), for download and reuse under an attributable, non-commercial creative commons license. Additionally, we have compiled a 'Landmarks Table' of key morphological features and marker gene expression that can be used to distinguish stages quickly and reliably (https://www.xenbase.org/entry/landmarks-table.do). This new open-access resource will facilitate Xenopus research and teaching in the decades to come.
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Bases de Datos Genéticas , Genómica , Animales , Humanos , Metamorfosis Biológica , Reproducibilidad de los Resultados , Xenopus laevis/genéticaRESUMEN
The swimming larvae of many marine animals identify a location on the seafloor to settle and undergo metamorphosis based on the presence of specific surface-bound bacteria. While bacteria-stimulated metamorphosis underpins processes such as the fouling of ship hulls, animal development in aquaculture, and the recruitment of new animals to coral reef ecosystems, little is known about the mechanisms governing this microbe-animal interaction. Here we review what is known and what we hope to learn about how bacteria and the factors they produce stimulate animal metamorphosis. With a few emerging model systems, including the tubeworm Hydroides elegans, corals, and the hydrozoan Hydractinia, we have begun to identify bacterial cues that stimulate animal metamorphosis and test hypotheses addressing their mechanisms of action. By understanding the mechanisms by which bacteria promote animal metamorphosis, we begin to illustrate how, and explore why, the developmental decision of metamorphosis relies on cues from environmental bacteria.
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Bacterias/metabolismo , Interacciones Microbiota-Huesped , Larva/microbiología , Metamorfosis Biológica , Poliquetos/crecimiento & desarrollo , Poliquetos/microbiología , Animales , Antozoos/microbiología , Organismos Acuáticos/microbiología , Bacterias/clasificación , Bacterias/genética , Arrecifes de Coral , EcosistemaRESUMEN
We used a representative of one of the oldest extant vertebrate lineages (jawless fish or agnathans) to investigate the early evolution and function of the growth hormone (GH)/prolactin (PRL) family. We identified a second member of the GH/PRL family in an agnathan, the sea lamprey (Petromyzon marinus). Structural, phylogenetic, and synteny analyses supported the identification of this hormone as prolactin-like (PRL-L), which has led to added insight into the evolution of the GH/PRL family. At least two ancestral genes were present in early vertebrates, which gave rise to distinct GH and PRL-L genes in lamprey. A series of gene duplications, gene losses, and chromosomal rearrangements account for the diversity of GH/PRL-family members in jawed vertebrates. Lamprey PRL-L is produced in the proximal pars distalis of the pituitary and is preferentially bound by the lamprey PRL receptor, whereas lamprey GH is preferentially bound by the lamprey GH receptor. Pituitary PRL-L messenger RNA (mRNA) levels were low in larvae, then increased significantly in mid-metamorphic transformers (stage 3); thereafter, levels subsided in final-stage transformers and metamorphosed juveniles. The abundance of PRL-L mRNA and immunoreactive protein increased in the pituitary of juveniles under hypoosmotic conditions, and treatment with PRL-L blocked seawater-associated inhibition of freshwater ion transporters. These findings clarify the origin and divergence of GH/PRL family genes in early vertebrates and reveal a function of PRL-L in osmoregulation of sea lamprey, comparable to a role of PRLs that is conserved in jawed vertebrates.
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Hormona de Crecimiento Humana , Petromyzon , Animales , Hormona del Crecimiento/genética , Hormona del Crecimiento/metabolismo , Osmorregulación/genética , Petromyzon/genética , Petromyzon/metabolismo , Filogenia , Prolactina/genética , Prolactina/metabolismo , ARN Mensajero/metabolismo , Vertebrados/genéticaRESUMEN
Juvenile hormones (JHs) control insect metamorphosis and reproduction. JHs act through a receptor complex consisting of methoprene-tolerant (Met) and taiman (Tai) proteins to induce transcription of specific genes. Among chemically diverse synthetic JH mimics (juvenoids), some of which serve as insecticides, unique peptidic juvenoids stand out as being highly potent yet exquisitely selective to a specific family of true bugs. Their mode of action is unknown. Here we demonstrate that, like established JH receptor agonists, peptidic juvenoids act upon the JHR Met to halt metamorphosis in larvae of the linden bug, Pyrrhocoris apterus. Peptidic juvenoids induced ligand-dependent dimerization between Met and Tai proteins from P. apterus but, consistent with their selectivity, not from other insects. A cell-based split-luciferase system revealed that the Met-Tai complex assembled within minutes of agonist presence. To explore the potential of juvenoid peptides, we synthesized 120 new derivatives and tested them in Met-Tai interaction assays. While many substituents led to loss of activity, improved derivatives active at sub-nanomolar range outperformed hitherto existing peptidic and classical juvenoids including fenoxycarb. Their potency in inducing Met-Tai interaction corresponded with the capacity to block metamorphosis in P. apterus larvae and to stimulate oogenesis in reproductively arrested adult females. Molecular modeling demonstrated that the high potency correlates with high affinity. This is a result of malleability of the ligand-binding pocket of P. apterus Met that allows larger peptidic ligands to maximize their contact surface. Our data establish peptidic juvenoids as highly potent and species-selective novel JHR agonists.
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Hormonas Juveniles , Metopreno , Animales , Femenino , Hormonas Juveniles/metabolismo , Ligandos , Metopreno/metabolismo , Insectos/metabolismo , Reproducción , Larva , Péptidos/farmacologíaRESUMEN
How larvae of the many phyla of marine invertebrates find places appropriate for settlement, metamorphosis, growth, and reproduction is an enduring question in marine science. Biofilm-induced metamorphosis has been observed in marine invertebrate larvae from nearly every major marine phylum. Despite the widespread nature of this phenomenon, the mechanism of induction remains poorly understood. The serpulid polychaete Hydroides elegans is a well established model for investigating bacteria-induced larval development. A broad range of biofilm bacterial species elicit larval metamorphosis in H. elegans via at least two mechanisms, including outer membrane vesicles (OMVs) and complexes of phage-tail bacteriocins. We investigated the interaction between larvae of H. elegans and the inductive bacterium Cellulophaga lytica, which produces an abundance of OMVs but not phage-tail bacteriocins. We asked whether the OMVs of C. lytica induce larval settlement due to cell membrane components or through delivery of specific cargo. Employing a biochemical structurefunction approach with a strong ecological focus, the cells and OMVs produced by C. lytica were interrogated to determine the class of the inductive compounds. Here, we report that larvae of H. elegans are induced to metamorphose by lipopolysaccharide produced by C. lytica. The widespread prevalence of lipopolysaccharide and its associated taxonomic and structural variability suggest it may be a broadly employed cue for bacterially induced larval settlement of marine invertebrates.
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Lipopolisacáridos , Metamorfosis Biológica , Animales , Bacterias , Biopelículas , Invertebrados/fisiología , Larva/fisiología , Lipopolisacáridos/farmacología , Metamorfosis Biológica/fisiologíaRESUMEN
Insects comprise over half of the described species, and the acquisition of metamorphosis must have contributed to their diversity and prosperity. The order Odonata (dragonflies and damselflies) is among the most-ancestral insects with drastic morphological changes upon metamorphosis, in which understanding of the molecular mechanisms will provide insight into the evolution of incomplete and complete metamorphosis in insects. In order to identify metamorphosis-related genes in Odonata, we performed comprehensive RNA-sequencing of the blue-tailed damselfly Ischnura senegalensis at different developmental stages. Comparative RNA-sequencing analyses between nymphs and adults identified eight nymph-specific and seven adult-specific transcripts. RNA interference (RNAi) of these candidate genes demonstrated that three transcription factors, Krüppel homolog 1 (Kr-h1), broad, and E93 play important roles in metamorphosis of both I. senegalensis and a phylogenetically distant dragonfly, Pseudothemis zonataE93 is essential for adult morphogenesis, and RNAi of Kr-h1 induced precocious metamorphosis in epidermis via up-regulation of E93 Precocious metamorphosis was also induced by RNAi of the juvenile hormone receptor Methoprene-tolerant (Met), confirming that the regulation of metamorphosis by the MEKRE93 (Met-Kr-h1-E93) pathway is conserved across diverse insects including the basal insect lineage Odonata. Notably, RNAi of broad produced unique grayish pigmentation on the nymphal abdominal epidermis. Survey of downstream genes for Kr-h1, broad, and E93 uncovered that unlike other insects, broad regulates a substantial number of nymph-specific and adult-specific genes independently of Kr-h1 and E93 These findings highlight the importance of functional changes and rewiring of the transcription factors Kr-h1, broad, and E93 in the evolution of insect metamorphosis.
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Evolución Biológica , Metamorfosis Biológica/genética , Odonata/crecimiento & desarrollo , Alas de Animales , Animales , Femenino , Perfilación de la Expresión Génica , Genes de Insecto , Masculino , Odonata/genética , Interferencia de ARNRESUMEN
BACKGROUND: Life cycle evolution includes ecological transitions and shifts in the timing of somatic and reproductive development (heterochrony). However, heterochronic changes can be tissue-specific, ultimately leading to the differential diversification of traits. Salamanders exhibit alternative life cycle polymorphisms involving either an aquatic to terrestrial metamorphosis (biphasic) or retention of aquatic larval traits into adulthood (paedomorphic). In this study, we used gene expression and histology to evaluate how life cycle evolution impacts temporal reproductive patterns in males of a polymorphic salamander. RESULTS: We found that heterochrony shifts the distribution of androgen signaling in the integument, which is correlated with significant differences in seasonal reproductive gland development and pheromone gene expression. In the testes, androgen receptor (ar) expression does not significantly vary between morphs or across seasons. We found significant differences in the onset of spermatogenesis, but by peak breeding season the testes were the same with respect to both histology and gene expression. CONCLUSION: This study provides an example of how seasonal heterochronic shifts in tissue-specific ar gene expression can disparately impact seasonal development and expression patterns across tissues, providing a potential mechanism for differential diversification of reproductive traits.