RESUMO
Insects have about 50 neuropeptide genes and about 70 genes, coding for neuropeptide G protein-coupled receptors (GPCRs). An important, but small family of evolutionarily related insect neuropeptides consists of adipokinetic hormone (AKH), corazonin, and AKH/corazonin-related peptide (ACP). Normally, insects have one specific GPCR for each of these neuropeptides. The tick Ixodes scapularis is not an insect, but belongs to the subphylum Chelicerata, which comprises ticks, scorpions, mites, spiders, and horseshoe crabs. Many of the neuropeptides and neuropeptide GPCRs occurring in insects, also occur in chelicerates, illustrating that insects and chelicerates are evolutionarily closely related. The tick I. scapularis is an ectoparasite and health risk for humans, because it infects its human host with dangerous pathogens during a blood meal. Understanding the biology of ticks will help researchers to prevent tick-borne diseases. By annotating the I. scapularis genome sequence, we previously found that ticks contain as many as five genes, coding for presumed ACP receptors. In the current paper, we cloned these receptors and expressed each of them in Chinese Hamster Ovary (CHO) cells. Each expressed receptor was activated by nanomolar concentrations of ACP, demonstrating that all five receptors were functional ACP receptors. Phylogenetic tree analyses showed that the cloned tick ACP receptors were mostly related to insect ACP receptors and, next, to insect AKH receptors, suggesting that ACP receptor genes and AKH receptor genes originated by gene duplications from a common ancestor. Similar duplications have probably occurred for the ligand genes, during a process of ligand/receptor co-evolution. Interestingly, chelicerates, in contrast to all other arthropods, do not have AKH or AKH receptor genes. Therefore, the ancestor of chelicerates might have lost AKH and AKH receptor genes and functionally replaced them by ACP and ACP receptor genes. For the small family of AKH, ACP, and corazonin receptors and their ligands, gene losses and gene gains occur frequently between the various ecdysozoan clades. Tardigrades, for example, which are well known for their survival in extreme environments, have as many as ten corazonin receptor genes and six corazonin peptide genes, while insects only have one of each, or none.
Assuntos
Hormônios de Inseto , Ixodes , Neuropeptídeos , Oligopeptídeos , Ácido Pirrolidonocarboxílico , Receptores Acoplados a Proteínas G , Animais , Neuropeptídeos/metabolismo , Neuropeptídeos/genética , Hormônios de Inseto/metabolismo , Hormônios de Inseto/genética , Ixodes/metabolismo , Ixodes/genética , Receptores Acoplados a Proteínas G/metabolismo , Receptores Acoplados a Proteínas G/genética , Oligopeptídeos/metabolismo , Oligopeptídeos/genética , Oligopeptídeos/química , Ácido Pirrolidonocarboxílico/análogos & derivados , Ácido Pirrolidonocarboxílico/metabolismo , Filogenia , Sequência de Aminoácidos , Cricetulus , Células CHO , Proteínas de Insetos/genética , Proteínas de Insetos/metabolismo , Receptores de Neuropeptídeos/metabolismo , Receptores de Neuropeptídeos/genéticaRESUMO
Many insects produce the cyclic neuropeptide inotocin (CLITNCPRGamide), which is the insect orthologue of the mammalian neuropeptides oxytocin and vasopressin. These insects also have one inotocin G protein-coupled receptor (GPCR), which is the orthologue of the mammalian oxytocin and vasopressin receptors. The tick Ixodes scapularis belongs to the subphylum Chelicerata, an arthropod taxon different from insects, to which also spiders, scorpions, and mites belong. I. scapularis is an ectoparasite and a health risk for humans, because it transfers pathogenic microorganisms to its human host during a blood meal, thereby causing serious neurological diseases, among them Lyme disease and tick-borne encephalitis (TBE). By annotating the genomic sequence of I. scapularis, we previously found one presumed tick inotocin preprohormone gene and, in contrast to insects, three genes coding for presumed inotocin GPCRs. We now find that these GPCR genes cluster on one genomic contig, suggesting that they originated by recent gene duplications. Closely located on the same contig are also four adipokinetic hormone/corazonin-related peptide (ACP) GPCR genes, and one crustacean cardioactive peptide (CCAP) GPCR gene, suggesting evolutionary relationships. These evolutionary relationships are confirmed by phylogenetic tree analyses of their gene products. We also cloned the tick inotocin preprohormone, which has a structural organization closely resembling mammalian oxytocin and vasopressin preprohormones, including the presence of a conserved neurophysin sequence, having seven cystine bridges. This neurophysin sequence has two cystine-knot domains, but in contrast to mammalian neurophysins, the tick neurophysin contains a canonical prohormone convertase cleavage signal and a peptide C-terminal amidation sequence (GKR), suggesting cleavage into two biologically active cystine-knot peptides. This cleavage/amidation sequence occurs in neurophysins from most hard tick species, but not in other chelicerates. Mature tick inotocin is different from insect inotocin and has the sequence CFITNCPPGamide. Finally, we cloned and stably expressed the three tick inotocin receptors in Chinese Hamster Ovary cells and found that each of them was activated by nanomolar concentrations of tick inotocin (EC50 for ITR1 = 1.6 × 10-8 M; EC50 for ITR2 = 5.8 × 10-9 M; EC50 for ITR3 = 9.3 × 10-9 M), thereby establishing that they are genuine tick inotocin receptors.
RESUMO
We have identified a corazonin G protein-coupled receptor (GPCR) gene in the tick Ixodes scapularis, which likely plays a central role in the physiology and behavior of this ectoparasite. This receptor gene is unusually large (1.133 Mb) and yields two corazonin (CRZ) receptor splice variants, where nearly half of the coding regions are exchanged: CRZ-Ra (containing exon 2, exon 3, and exon 4 of the gene) and CRZ-Rb (containing exon 1, exon 3, and exon 4 of the gene). CRZ-Ra codes for a GPCR with a canonical DRF sequence at the border of the third transmembrane helix and the second intracellular loop. The positively-charged R residue from the DRF sequence is important for coupling of G proteins after activation of a GPCR. CRZ-Rb, in contrast, codes for a GPCR with an unusual DQL sequence at this position, still retaining a negatively-charged D residue, but lacking a positively-charged R residue, suggesting different G protein coupling. Another difference between the two splice variants is that exon 2 from CRZ-Ra codes for an N-terminal signal sequence. Normally, GPCRs do not have N-terminal signal sequences, although a few mammalian GPCRs have. In the tick CRZ-Ra, the signal sequence probably assists with inserting the receptor correctly into the RER membrane. We stably transfected Chinese Hamster Ovary cells with each of the two splice variants and carried out bioluminescence bioassays that also included the use of the human promiscuous G protein G16. CRZ-Ra turned out to be selective for I. scapularis corazonin (EC50 = 10-8 M) and could not be activated by related neuropeptides like adipokinetic hormone (AKH) and AKH/corazonin-related peptide (ACP). Similarly, also CRZ-Rb could only be activated by corazonin, although about 4-fold higher concentrations were needed to activate it (EC50 = 4 x 10-8 M). The genomic organization of the tick corazonin GPCR gene is similar to that of the insect AKH and ACP receptor genes. This similar genomic organization can also be found in the human gonadotropin-releasing hormone (GnRH) receptor gene, confirming previous conclusions that the corazonin, AKH, and ACP receptor genes are the true arthropod orthologues of the human GnRH receptor gene.
Assuntos
Ixodes , Neuropeptídeos , Animais , Cricetinae , Humanos , Ixodes/genética , Ixodes/metabolismo , Células CHO , Cricetulus , Neuropeptídeos/genética , Proteínas de Insetos/genética , Receptores Acoplados a Proteínas G/genética , Sinais Direcionadores de ProteínasRESUMO
Bilateria have bilateral symmetry and are subdivided into Deuterostomia (animals like vertebrates) and Protostomia (animals like insects and mollusks). Neuropeptides occur in both Proto- and Deuterostomia and they are frequently structurally related across these two lineages. For example, peptides belonging to the oxytocin/vasopressin family exist in both clades. The same is true for the G protein-coupled receptors (GPCRs) of these peptides. These observations suggest that these neuropeptides and their GPCRs were already present in the common ancestor of Proto- and Deuterostomia, which lived about 700 million years ago (MYA). Furthermore, neuropeptides and their GPCRs occur in two early-branching phyla that diverged before the emergence of Bilateria: Cnidaria (animals like corals and sea anemones), and Placozoa (small disk-like animals, feeding on algae). The sequences of these neuropeptides and their GPCRs, however, are not closely related to those from Bilateria. In addition, cnidarian neuropeptides and their receptors are not closely related to those from Placozoa. We propose that the divergence times between Cnidaria, Placozoa, and Bilateria might be too long for recognizing sequence identities. Leucine-rich repeats-containing GPCRs (LGRs) are a special class of GPCRs that are characterized by a long N-terminus containing 10-20 leucine-rich domains, which are used for ligand binding. Among the ligands for LGRs are dimeric glycoprotein hormones, and insulin-like peptides, such as relaxin. LGRs have been found not only in Proto- and Deuterostomia, but also in early emerging phyla, such as Cnidaria and Placozoa. Humans have eight LGRs. In our current review, we have revisited the annotations of LGRs from the sea anemone Nematostella vectensis and the placozoan Trichoplax adhaerens. We identified 13 sea anemone LGRs and no less than 46 LGRs from T. adhaerens. All eight human LGRs appear to have orthologues in sea anemones and placozoans. LGRs and their ligands, therefore, have a long evolutionary history, going back to the common ancestor of Cnidaria and Placozoa.
Assuntos
Insulinas , Neuropeptídeos , Placozoa , Relaxina , Anêmonas-do-Mar , Animais , Glicoproteínas/metabolismo , Humanos , Leucina , Ligantes , Neuropeptídeos/genética , Neuropeptídeos/metabolismo , Ocitocina/metabolismo , Placozoa/genética , Placozoa/metabolismo , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Relaxina/metabolismoRESUMO
BACKGROUND: The animal phylum Cnidaria consists of six classes or subphyla: Hydrozoa, Scyphozoa, Cubozoa, Staurozoa, Anthozoa, and Endocnidozoa. Cnidarians have an early evolutionary origin, diverging before the emergence of the Bilateria. Extant members from this phylum, therefore, are important resources for understanding the evolution of the nervous system. Cnidarian nervous systems are strongly peptidergic. Using genomics, we have recently shown that three neuropeptide families (the X1PRX2amides, GRFamides, and GLWamides) are wide-spread in four (Scyphozoa, Cubozoa, Staurozoa, Anthozoa) out of six cnidarian classes or subphyla, suggesting that these three neuropeptide families emerged in the common cnidarian ancestor. In the current paper, we analyze the remaining cnidarian class, Hydrozoa, and the subphylum Endocnidozoa, to make firm conclusions about the evolution of neuropeptide genes in Cnidaria. RESULTS: We analyzed sixteen hydrozoan species with a sequenced genome or transcriptome, using a recently developed software program for discovering neuropeptide genes. These species belonged to various hydrozoan subclasses and orders, among them the laboratory models Hydra, Hydractinia, and Clytia. We found that each species contained three to five neuropeptide families. A common feature for all hydrozoans was that they contained genes coding for (i) X1PRX2amide peptides, (ii) GRFamide peptides, and (iii) GLWamide peptides. These results support our previous conclusions that these three neuropeptide families evolved early in evolution. In addition to these three neuropeptide families, hydrozoans expressed up to two other neuropeptide gene families, which, however, were only occurring in certain animal groups. Endocnidozoa (Myxozoa) are microscopically small endoparasites, which are strongly reduced. For long, it was unknown to which phylum these parasites belonged, but recently they have been associated with cnidarians. We analyzed nine endocnidozoan species and found that two of them (Polypodium hydriforme and Buddenbrockia plumatellae) expressed neuropeptide genes. These genes coded for neuropeptides belonging to the GRFamide and GLWamide families with structures closely resembling them from hydrozoans. CONCLUSIONS: We found X1PRX2amide, GRFamide, and GLWamide peptides in all species belonging to the Hydrozoa, confirming that these peptides originated in the common cnidarian ancestor. In addition, we discovered GRFamide and GLWamide peptide genes in some members of the Endocnidozoa, thereby linking these parasites to Hydrozoa.
Assuntos
Cnidários , Hidrozoários , Myxozoa , Neuropeptídeos , Animais , Cnidários/genética , Evolução Molecular , Genômica , Hidrozoários/genética , Myxozoa/genética , Neuropeptídeos/genética , FilogeniaRESUMO
Box jellyfish have an elaborate visual system and perform advanced visually guided behaviors. However, the rhopalial nervous system (RNS), believed to be the main visual processing center, only has 1000 neurons in each of the four eye carrying rhopalia. We have examined the detailed structure of the RNS of the box jellyfish Tripedalia cystophora, using immunolabeling with antibodies raised against four putative neuropeptides (T. cystophora RFamide, VWamide, RAamide, and FRamide). In the RNS, T. cystophora RF-, VW-, and RAamide antibodies stain sensory neurons, the pit eyes, the neuropil, and peptide-specific subpopulations of stalk-associated neurons and giant neurons. Furthermore, RFamide ir+ neurites are seen in the epidermal stalk nerve, whereas VWamide antibodies stain the gastrodermal stalk nerve. RFamide has the most widespread expression including in the ring and radial nerves, the pedalium nerve plexus, and the tentacular nerve net. RAamide is the putative neurotransmitter in the motor neurons of the subumbrellar nerve net, and VWamide is a potential marker for neuronal differentiation as it is found in subpopulations of undifferentiated cells both in the rhopalia and in the bell. The results from the FRamide antibodies were not included as only few cells were stained, and in an unreproducible way. Our studies show hitherto-unseen details of the nervous system of T. cystophora and allowed us to identify specific functional groups of neurons. This identification is important for understanding visual processing in the RNS and enables experimental work, directly addressing the role of the different neuropeptides in vision.
Assuntos
Cubomedusas/metabolismo , Rede Nervosa/metabolismo , Neuropeptídeos/biossíntese , Neurópilo/metabolismo , Vias Visuais/metabolismo , Fatores Etários , Animais , Cubomedusas/química , Cubomedusas/genética , Expressão Gênica , Rede Nervosa/química , Sistema Nervoso/química , Sistema Nervoso/metabolismo , Neuritos/química , Neuritos/metabolismo , Neuropeptídeos/análise , Neuropeptídeos/genética , Neurópilo/química , Células Receptoras Sensoriais/química , Células Receptoras Sensoriais/metabolismo , Vias Visuais/químicaRESUMO
BACKGROUND: Nervous systems originated before the split of Proto- and Deuterostomia, more than 600 million years ago. Four animal phyla (Cnidaria, Placozoa, Ctenophora, Porifera) diverged before this split and studying these phyla could give us important information on the evolution of the nervous system. Here, we have annotated the neuropeptide preprohormone genes of twenty species belonging to the subclass Hexacorallia or Ceriantharia (Anthozoa: Cnidaria), using thirty-seven publicly accessible genome or transcriptome databases. Studying hexacorals is important, because they are versatile laboratory models for development (e.g., Nematostella vectensis) and symbiosis (e.g., Exaiptasia diaphana) and also are prominent reef-builders. RESULTS: We found that each hexacoral or ceriantharian species contains five to ten neuropeptide preprohormone genes. Many of these preprohormones contain multiple copies of immature neuropeptides, which can be up to 50 copies of identical or similar neuropeptide sequences. We also discovered preprohormones that only contained one neuropeptide sequence positioned directly after the signal sequence. Examples of them are neuropeptides that terminate with the sequence RWamide (the Antho-RWamides). Most neuropeptide sequences are N-terminally protected by pyroglutamyl (pQ) or one or more prolyl residues, while they are C-terminally protected by an amide group. Previously, we isolated and sequenced small neuropeptides from hexacorals that were N-terminally protected by an unusual L-3-phenyllactyl group. In our current analysis, we found that these N-phenyllactyl-peptides are derived from N-phenylalanyl-peptides located directly after the signal sequence of the preprohormone. The N-phenyllactyl- peptides appear to be confined to the hexacorallian order Actiniaria and do not occur in other cnidarians. On the other hand, (1) the neuropeptide Antho-RFamide (pQGRFamide); (2) peptides with the C-terminal sequence GLWamide; and (3) tetrapeptides with the X1PRX2amide consensus sequence (most frequently GPRGamide) are ubiquitous in Hexacorallia. CONCLUSIONS: We found GRFamide, GLWamide, and X1PRX2amide peptides in all tested Hexacorallia. Previously, we discovered these three neuropeptide classes also in Cubozoa, Scyphozoa, and Staurozoa, indicating that these neuropeptides originated in the common cnidarian ancestor and are evolutionarily ancient. In addition to these ubiquitous neuropeptides, other neuropeptides appear to be confined to specific cnidarian orders or subclasses.
Assuntos
Neuropeptídeos/genética , Anêmonas-do-Mar/genética , Motivos de Aminoácidos , Animais , Família Multigênica , Neuropeptídeos/química , Filogenia , Precursores de Proteínas/química , Precursores de Proteínas/genética , Anêmonas-do-Mar/classificação , TranscriptomaRESUMO
The tremendous diversity of Hymenoptera is commonly attributed to the evolution of parasitoidism in the last common ancestor of parasitoid sawflies (Orussidae) and wasp-waisted Hymenoptera (Apocrita). However, Apocrita and Orussidae differ dramatically in their species richness, indicating that the diversification of Apocrita was promoted by additional traits. These traits have remained elusive due to a paucity of sawfly genome sequences, in particular those of parasitoid sawflies. Here, we present comparative analyses of draft genomes of the primarily phytophagous sawfly Athalia rosae and the parasitoid sawfly Orussus abietinus. Our analyses revealed that the ancestral hymenopteran genome exhibited traits that were previously considered unique to eusocial Apocrita (e.g., low transposable element content and activity) and a wider gene repertoire than previously thought (e.g., genes for CO2 detection). Moreover, we discovered that Apocrita evolved a significantly larger array of odorant receptors than sawflies, which could be relevant to the remarkable diversification of Apocrita by enabling efficient detection and reliable identification of hosts.
Assuntos
Especiação Genética , Genoma de Inseto , Interações Hospedeiro-Parasita/genética , Himenópteros/genética , Sequência de Aminoácidos , Animais , Sequência Conservada , Elementos de DNA Transponíveis , Feminino , Dosagem de Genes , Glicoproteínas/genética , Herbivoria/genética , Imunidade/genética , Proteínas de Insetos/genética , Masculino , Família Multigênica , Receptores Odorantes/genética , Comportamento Social , Visão Ocular/genéticaRESUMO
During animal evolution, ancestral Cnidaria and Bilateria diverged more than 600 million years ago. The nervous systems of extant cnidarians are strongly peptidergic. Neuropeptides have been isolated and sequenced from a few model cnidarians, but a global investigation of the presence of neuropeptides in all cnidarian classes has been lacking. Here, we have used a recently developed software program to annotate neuropeptides in the publicly available genomes and transcriptomes from members of the classes Cubozoa, Scyphozoa, and Staurozoa (which all belong to the subphylum Medusozoa) and contrasted these results with neuropeptides present in the subclass Octocorallia (belonging to the class Anthozoa). We found three to six neuropeptide preprohormone genes in members of the above-mentioned cnidarian classes or subclasses, each coding for several (up to thirty-two) similar or identical neuropeptide copies. Two of these neuropeptide preprohormone genes are present in all cnidarian classes/subclasses investigated, so they are good candidates for being among the first neuropeptide genes evolved in cnidarians. One of these primordial neuropeptide genes codes for neuropeptides having the C-terminal sequence GRFamide (pQGRFamide in Octocorallia; pQWLRGRFamide in Cubozoa and Scyphozoa; pQFLRGRFamide in Staurozoa). The other primordial neuropeptide gene codes for peptides having RPRSamide or closely resembling amino acid sequences. In addition to these two primordial neuropeptide sequences, cnidarians have their own class- or subclass-specific neuropeptides, which probably evolved to serve class/subclass-specific needs. When we carried out phylogenetic tree analyses of the GRFamide or RPRSamide preprohormones from cubozoans, scyphozoans, staurozoans, and octocorallia, we found that their phylogenetic relationships perfectly agreed with current models of the phylogeny of the studied cnidarian classes and subclasses. These results support the early origins of the GRFamide and RPRSamide preprohormone genes.
RESUMO
BACKGROUND: The phyla Cnidaria, Placozoa, Ctenophora, and Porifera emerged before the split of proto- and deuterostome animals, about 600 million years ago. These early metazoans are interesting, because they can give us important information on the evolution of various tissues and organs, such as eyes and the nervous system. Generally, cnidarians have simple nervous systems, which use neuropeptides for their neurotransmission, but some cnidarian medusae belonging to the class Cubozoa (box jellyfishes) have advanced image-forming eyes, probably associated with a complex innervation. Here, we describe a new transcriptome database from the cubomedusa Tripedalia cystophora. RESULTS: Based on the combined use of the Illumina and PacBio sequencing technologies, we produced a highly contiguous transcriptome database from T. cystophora. We then developed a software program to discover neuropeptide preprohormones in this database. This script enabled us to annotate seven novel T. cystophora neuropeptide preprohormone cDNAs: One coding for 19 copies of a peptide with the structure pQWLRGRFamide; one coding for six copies of a different RFamide peptide; one coding for six copies of pQPPGVWamide; one coding for eight different neuropeptide copies with the C-terminal LWamide sequence; one coding for thirteen copies of a peptide with the RPRAamide C-terminus; one coding for four copies of a peptide with the C-terminal GRYamide sequence; and one coding for seven copies of a cyclic peptide, of which the most frequent one has the sequence CTGQMCWFRamide. We could also identify orthologs of these seven preprohormones in the cubozoans Alatina alata, Carybdea xaymacana, Chironex fleckeri, and Chiropsalmus quadrumanus. Furthermore, using TBLASTN screening, we could annotate four bursicon-like glycoprotein hormone subunits, five opsins, and 52 other family-A G protein-coupled receptors (GPCRs), which also included two leucine-rich repeats containing G protein-coupled receptors (LGRs) in T. cystophora. The two LGRs are potential receptors for the glycoprotein hormones, while the other GPCRs are candidate receptors for the above-mentioned neuropeptides. CONCLUSIONS: By combining Illumina and PacBio sequencing technologies, we have produced a new high-quality de novo transcriptome assembly from T. cystophora that should be a valuable resource for identifying the neuronal components that are involved in vision and other behaviors in cubomedusae.
Assuntos
Cubomedusas/genética , Peptídeos/genética , Transmissão Sináptica/genética , Transcriptoma/genética , Animais , Cubomedusas/fisiologia , Humanos , Neurônios/metabolismo , Neuropeptídeos , Opsinas/genética , Receptores Acoplados a Proteínas G/genética , Visão Ocular/genética , Visão Ocular/fisiologiaRESUMO
Most multicellular animals belong to two evolutionary lineages, the Proto- and Deuterostomia, which diverged 640-760 million years (MYR) ago. Neuropeptide signaling is abundant in animals belonging to both lineages, but it is often unclear whether there exist evolutionary relationships between the neuropeptide systems used by proto- or deuterostomes. An exception, however, are members of the gonadotropin-releasing hormone (GnRH) receptor superfamily, which occur in both evolutionary lineages, where GnRHs are the ligands in Deuterostomia and GnRH-like peptides, adipokinetic hormone (AKH), corazonin, and AKH/corazonin-related peptide (ACP) are the ligands in Protostomia. AKH is a well-studied insect neuropeptide that mobilizes lipids and carbohydrates from the insect fat body during flight. In our present paper, we show that AKH is not only widespread in insects, but also in other Ecdysozoa and in Lophotrochozoa. Furthermore, we have cloned and deorphanized two G protein-coupled receptors (GPCRs) from the oyster Crassostrea gigas (Mollusca) that are activated by low nanomolar concentrations of oyster AKH (pQVSFSTNWGSamide). Our discovery of functional AKH receptors in molluscs is especially significant, because it traces the emergence of AKH signaling back to about 550 MYR ago and brings us closer to a more complete understanding of the evolutionary origins of the GnRH receptor superfamily.
Assuntos
Adipocinas/metabolismo , Evolução Biológica , Hormônios de Inseto/metabolismo , Invertebrados/metabolismo , Oligopeptídeos/metabolismo , Ácido Pirrolidonocarboxílico/análogos & derivados , Receptores Acoplados a Proteínas G/metabolismo , Animais , Células CHO , Clonagem Molecular , Biologia Computacional , Crassostrea/metabolismo , Cricetinae , Cricetulus , Drosophila melanogaster , Hormônio Liberador de Gonadotropina/metabolismo , Humanos , Proteínas de Insetos/metabolismo , Insetos , Ligantes , Neuropeptídeos/metabolismo , Peptídeos/metabolismo , Filogenia , Ácido Pirrolidonocarboxílico/metabolismo , Transdução de SinaisRESUMO
The neuroendocrine peptides CCHamide-1 and -2, encoded by the genes ccha1 and -2, are produced by endocrine cells in the midgut and by neurons in the brain of Drosophila melanogaster. Here, we used the CRISPR/Cas9 technique to disrupt the ccha1 and -2 genes and identify mutant phenotypes with a focus on ccha-2 mutants. We found that both larval and adult ccha2 mutants showed a significantly reduced food intake as measured in adult flies by the Capillary Feeding (CAFE) assay (up to 72% reduced food intake compared to wild-type). Locomotion tests in adult flies showed that ccha2 mutants had a significantly reduced locomotor activity especially around 8 a.m. and 8 p.m., where adult Drosophila normally feeds (up to 70% reduced locomotor activity compared to wild-type). Reduced larval feeding is normally coupled to a delayed larval development, a process that is mediated by insulin. Accordingly, we found that the ccha2 mutants had a remarkably delayed development, showing pupariation 70 hours after the pupariation time point of the wild-type. In contrast, the ccha-1 mutants were not developmentally delayed. We also found that the ccha2 mutants had up to 80% reduced mRNA concentrations coding for the Drosophila insulin-like-peptides-2 and -3, while these concentrations were unchanged for the ccha1 mutants. From these experiments we conclude that CCHamide-2 is an orexigenic peptide and an important factor for controlling developmental timing in Drosophila.
Assuntos
Encéfalo/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila/metabolismo , Mucosa Intestinal/metabolismo , Neuropeptídeos/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , DNA , Drosophila/crescimento & desenvolvimento , Drosophila/fisiologia , Proteínas de Drosophila/genética , Comportamento Alimentar , Larva/metabolismo , Locomoção , Dados de Sequência Molecular , Mutação , Neuropeptídeos/genéticaRESUMO
Muscarinic acetylcholine receptors (mAChRs) are G protein-coupled receptors (GPCRs) that are activated by the agonists acetylcholine and muscarine and blocked by several antagonists, among them atropine. In mammals five mAChRs (m1-m5) exist of which m1, m3, and m5 are coupled to members of the Gq/11 family and m2 and m4 to members of the Gi/0 family. We have recently shown that Drosophila melanogaster and other arthropods have two mAChRs, named A and B, where the A-type has the same pharmacology as the mammalian mAChRs, while the B-type has a very low affinity to muscarine and no affinity to classical antagonists such as atropine. Here, we find that the D. melanogaster A-type mAChR is coupled to Gq/11 and D. melanogaster B-type mAChR to Gi/0. Furthermore, by comparing the second and third intracellular loops of all animal mAChRs for which the G protein coupling has been established, we could identify several amino acid residues likely to be specific for either Gq/11 or Gi/0 coupling. Using these hallmarks for specific mAChR G protein interaction we found that all protostomes with a sequenced genome have one mAChR coupled to Gq/11 and one to four mAChRs coupled to Gi/0. Furthermore, in protostomes, probably all A-type mAChRs are coupled to Gq/11 and all B-type mAChRs to G0/i.
Assuntos
Isoformas de Proteínas/metabolismo , Receptores Muscarínicos/metabolismo , Sistemas do Segundo Mensageiro , Sequência de Aminoácidos , Animais , Células CHO , Cricetinae , Cricetulus , Drosophila melanogaster , Dados de Sequência Molecular , Isoformas de Proteínas/química , Receptores Muscarínicos/química , Homologia de Sequência de AminoácidosRESUMO
The evolution of eusociality is one of the major transitions in evolution, but the underlying genomic changes are unknown. We compared the genomes of 10 bee species that vary in social complexity, representing multiple independent transitions in social evolution, and report three major findings. First, many important genes show evidence of neutral evolution as a consequence of relaxed selection with increasing social complexity. Second, there is no single road map to eusociality; independent evolutionary transitions in sociality have independent genetic underpinnings. Third, though clearly independent in detail, these transitions do have similar general features, including an increase in constrained protein evolution accompanied by increases in the potential for gene regulation and decreases in diversity and abundance of transposable elements. Eusociality may arise through different mechanisms each time, but would likely always involve an increase in the complexity of gene networks.
Assuntos
Abelhas/genética , Evolução Molecular , Deriva Genética , Comportamento Social , Transcriptoma , Aminoácido N-Acetiltransferase , Animais , Abelhas/classificação , Elementos de DNA Transponíveis , Regulação da Expressão Gênica , Redes Reguladoras de Genes , Genoma de Inseto/genética , Filogenia , Seleção Genética , Fatores de Transcrição/química , Fatores de Transcrição/genéticaRESUMO
BACKGROUND: The shift from solitary to social behavior is one of the major evolutionary transitions. Primitively eusocial bumblebees are uniquely placed to illuminate the evolution of highly eusocial insect societies. Bumblebees are also invaluable natural and agricultural pollinators, and there is widespread concern over recent population declines in some species. High-quality genomic data will inform key aspects of bumblebee biology, including susceptibility to implicated population viability threats. RESULTS: We report the high quality draft genome sequences of Bombus terrestris and Bombus impatiens, two ecologically dominant bumblebees and widely utilized study species. Comparing these new genomes to those of the highly eusocial honeybee Apis mellifera and other Hymenoptera, we identify deeply conserved similarities, as well as novelties key to the biology of these organisms. Some honeybee genome features thought to underpin advanced eusociality are also present in bumblebees, indicating an earlier evolution in the bee lineage. Xenobiotic detoxification and immune genes are similarly depauperate in bumblebees and honeybees, and multiple categories of genes linked to social organization, including development and behavior, show high conservation. Key differences identified include a bias in bumblebee chemoreception towards gustation from olfaction, and striking differences in microRNAs, potentially responsible for gene regulation underlying social and other traits. CONCLUSIONS: These two bumblebee genomes provide a foundation for post-genomic research on these key pollinators and insect societies. Overall, gene repertoires suggest that the route to advanced eusociality in bees was mediated by many small changes in many genes and processes, and not by notable expansion or depauperation.
Assuntos
Abelhas/genética , Comportamento Animal , Genes de Insetos , Comportamento Social , Animais , Venenos de Abelha/genética , Abelhas/classificação , Abelhas/fisiologia , Células Quimiorreceptoras/metabolismo , Mapeamento Cromossômico , Bases de Dados Genéticas , Evolução Molecular , Feminino , Regulação da Expressão Gênica , Rearranjo Gênico , Genômica , Sequências Repetitivas Dispersas , Masculino , Fases de Leitura Aberta , Polimorfismo de Nucleotídeo Único , Selenoproteínas/genética , Selenoproteínas/metabolismo , Análise de Sequência de DNA , Especificidade da Espécie , SinteniaRESUMO
Gall-forming arthropods are highly specialized herbivores that, in combination with their hosts, produce extended phenotypes with unique morphologies [1]. Many are economically important, and others have improved our understanding of ecology and adaptive radiation [2]. However, the mechanisms that these arthropods use to induce plant galls are poorly understood. We sequenced the genome of the Hessian fly (Mayetiola destructor; Diptera: Cecidomyiidae), a plant parasitic gall midge and a pest of wheat (Triticum spp.), with the aim of identifying genic modifications that contribute to its plant-parasitic lifestyle. Among several adaptive modifications, we discovered an expansive reservoir of potential effector proteins. Nearly 5% of the 20,163 predicted gene models matched putative effector gene transcripts present in the M. destructor larval salivary gland. Another 466 putative effectors were discovered among the genes that have no sequence similarities in other organisms. The largest known arthropod gene family (family SSGP-71) was also discovered within the effector reservoir. SSGP-71 proteins lack sequence homologies to other proteins, but their structures resemble both ubiquitin E3 ligases in plants and E3-ligase-mimicking effectors in plant pathogenic bacteria. SSGP-71 proteins and wheat Skp proteins interact in vivo. Mutations in different SSGP-71 genes avoid the effector-triggered immunity that is directed by the wheat resistance genes H6 and H9. Results point to effectors as the agents responsible for arthropod-induced plant gall formation.
Assuntos
Cromossomos/genética , Dípteros/genética , Família Multigênica/genética , Filogenia , Tumores de Planta/genética , Triticum/parasitologia , Adaptação Biológica/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Dípteros/metabolismo , Larva/metabolismo , Modelos Genéticos , Dados de Sequência Molecular , Análise de Sequência de DNA , Homologia de Sequência , Comportamento Sexual Animal/fisiologia , Técnicas do Sistema de Duplo-Híbrido , Ubiquitina-Proteína Ligases/genéticaRESUMO
Myriapods (e.g., centipedes and millipedes) display a simple homonomous body plan relative to other arthropods. All members of the class are terrestrial, but they attained terrestriality independently of insects. Myriapoda is the only arthropod class not represented by a sequenced genome. We present an analysis of the genome of the centipede Strigamia maritima. It retains a compact genome that has undergone less gene loss and shuffling than previously sequenced arthropods, and many orthologues of genes conserved from the bilaterian ancestor that have been lost in insects. Our analysis locates many genes in conserved macro-synteny contexts, and many small-scale examples of gene clustering. We describe several examples where S. maritima shows different solutions from insects to similar problems. The insect olfactory receptor gene family is absent from S. maritima, and olfaction in air is likely effected by expansion of other receptor gene families. For some genes S. maritima has evolved paralogues to generate coding sequence diversity, where insects use alternate splicing. This is most striking for the Dscam gene, which in Drosophila generates more than 100,000 alternate splice forms, but in S. maritima is encoded by over 100 paralogues. We see an intriguing linkage between the absence of any known photosensory proteins in a blind organism and the additional absence of canonical circadian clock genes. The phylogenetic position of myriapods allows us to identify where in arthropod phylogeny several particular molecular mechanisms and traits emerged. For example, we conclude that juvenile hormone signalling evolved with the emergence of the exoskeleton in the arthropods and that RR-1 containing cuticle proteins evolved in the lineage leading to Mandibulata. We also identify when various gene expansions and losses occurred. The genome of S. maritima offers us a unique glimpse into the ancestral arthropod genome, while also displaying many adaptations to its specific life history.
Assuntos
Artrópodes/genética , Genoma , Sintenia , Animais , Peptídeos e Proteínas de Sinalização do Ritmo Circadiano/genética , Metilação de DNA , Evolução Molecular , Feminino , Genoma Mitocondrial , Hormônios/genética , Masculino , Família Multigênica , Filogenia , Polimorfismo Genético , Proteínas Quinases/genética , RNA não Traduzido/genética , Receptores Odorantes/genética , Selenoproteínas/genética , Cromossomos Sexuais , Fatores de Transcrição/genéticaRESUMO
Termites normally rely on gut symbionts to decompose organic matter but the Macrotermitinae domesticated Termitomyces fungi to produce their own food. This transition was accompanied by a shift in the composition of the gut microbiota, but the complementary roles of these bacteria in the symbiosis have remained enigmatic. We obtained high-quality annotated draft genomes of the termite Macrotermes natalensis, its Termitomyces symbiont, and gut metagenomes from workers, soldiers, and a queen. We show that members from 111 of the 128 known glycoside hydrolase families are represented in the symbiosis, that Termitomyces has the genomic capacity to handle complex carbohydrates, and that worker gut microbes primarily contribute enzymes for final digestion of oligosaccharides. This apparent division of labor is consistent with the Macrotermes gut microbes being most important during the second passage of comb material through the termite gut, after a first gut passage where the crude plant substrate is inoculated with Termitomyces asexual spores so that initial fungal growth and polysaccharide decomposition can proceed with high efficiency. Complex conversion of biomass in termite mounds thus appears to be mainly accomplished by complementary cooperation between a domesticated fungal monoculture and a specialized bacterial community. In sharp contrast, the gut microbiota of the queen had highly reduced plant decomposition potential, suggesting that mature reproductives digest fungal material provided by workers rather than plant substrate.
Assuntos
Isópteros/metabolismo , Plantas/metabolismo , Simbiose , Termitomyces/metabolismo , Animais , Bactérias/classificação , Bactérias/genética , Bactérias/metabolismo , Metabolismo dos Carboidratos , Sistema Digestório/metabolismo , Sistema Digestório/microbiologia , Feminino , Proteínas Fúngicas/metabolismo , Glicosídeo Hidrolases/metabolismo , Interações Hospedeiro-Patógeno , Isópteros/genética , Isópteros/microbiologia , Masculino , Metagenoma/genética , Consórcios Microbianos/genética , Consórcios Microbianos/fisiologia , Oligossacarídeos/metabolismo , Polissacarídeos/metabolismo , Análise de Sequência de DNA , Termitomyces/genética , Termitomyces/fisiologiaRESUMO
In this review we trace the evolutionary connections between GnRH receptors from vertebrates and the receptors for adipokinetic hormone (AKH), AKH/corazonin-related peptide (ACP), and corazonin from arthropods. We conclude that these G protein-coupled receptors (GPCRs) are closely related and have a common evolutionary origin, which dates back to the split of Proto- and Deuterostomia, about 700 million years ago. We propose that in the protostomian lineage, the ancestral GnRH-like receptor gene duplicated as did its GnRH-like ligand gene, followed by diversification, leading to (i) a corazonin receptor gene and a corazonin-like ligand gene, and (ii) an AKH receptor gene and an AKH-like ligand gene in the Mollusca and Annelida. Subsequently, the AKH receptor and ligand genes duplicated once more, yielding the situation that we know from arthropods today, where three independent hormonal systems exist, signalling with AKH, ACP, and corazonin. Our model for the evolution of GnRH signaling in the Protostomia is a striking example of receptor-ligand co-evolution. This model has been developed using several bioinformatics tools (TBLASTN searches, phylogenetic tree analyses), which also helped us to annotate six novel AKH preprohormones and their corresponding AKH sequences from the following molluscs: the sea hare Aplysia californica (AKH sequence: pQIHFSPDWGTamide), the sea slug Tritonia diomedea (pQIHFSPGWEPamide), the fresh water snail Bithynia siamensis goniomphalos (pQIHFTPGWGSamide), the owl limpet Lottia gigantea (pQIHFSPTWGSamide), the oyster Crassostrea gigas (pQVSFSTNWGSamide), and the freshwater pearl mussel Hyriopsis cumingii (pQISFSTNWGSamide). We also found AKHs in the tardigrade Hysibius dujardini (pQLSFTGWGHamide), the rotifer Brachionus calycifloros (pQLTFSSDWSGamide), and the penis worm Priapulus caudatus (pQIFFSKGWRGamide). This is the first report, showing that AKH signaling is widespread in molluscs.
Assuntos
Evolução Molecular , Hormônio Liberador de Gonadotropina/genética , Hormônios de Inseto/genética , Proteínas de Insetos/genética , Invertebrados/genética , Neuropeptídeos/genética , Oligopeptídeos/genética , Ácido Pirrolidonocarboxílico/análogos & derivados , Receptores LHRH/genética , Receptores de Neuropeptídeos/genética , Sequência de Aminoácidos , Animais , Hormônios de Inseto/metabolismo , Proteínas de Insetos/metabolismo , Ligantes , Masculino , Dados de Sequência Molecular , Família Multigênica/genética , Neuropeptídeos/metabolismo , Oligopeptídeos/metabolismo , Filogenia , Ácido Pirrolidonocarboxílico/metabolismo , Receptores Acoplados a Proteínas G/genética , Receptores de Neuropeptídeos/metabolismo , Homologia de Sequência de AminoácidosRESUMO
Neuropeptides are a highly diverse group of signaling molecules that affect a broad range of biological processes in insects, including development, metabolism, behavior, and reproduction. In the central nervous system, neuropeptides are usually considered to act as neuromodulators and cotransmitters that modify the effect of "classical" transmitters at the synapse. The present study analyzes the neuropeptide repertoire of higher cerebral neuropils in the brain of the red flour beetle Tribolium castaneum. We focus on two integrative neuropils of the olfactory pathway, the antennal lobes and the mushroom bodies. Using the technique of direct peptide profiling by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry, we demonstrate that these neuropils can be characterized by their specific neuropeptide expression profiles. Complementary immunohistological analyses of selected neuropeptides revealed neuropeptide distribution patterns within the antennal lobes and the mushroom bodies. Both approaches revealed consistent differences between the neuropils, underlining that direct peptide profiling by mass spectrometry is a fast and reliable method to identify neuropeptide content.