Ancient Arthropods Research Articles

A Crayfish Insulin-like-binding Protein: ANOTHER PIECE IN THE ANDROGENIC GLAND INSULIN-LIKE HORMONE PUZZLE IS REVEALED

Across the animal kingdom, the involvement of insulin-like peptide (ILP) signaling in sex-related differentiation processes is attracting increasing attention. Recently, a gender-specific ILP was identified as the androgenic sex hormone in Crustacea. However, moieties modulating the actions of this androgenic insulin-like growth factor were yet to be revealed. Through molecular screening of an androgenic gland (AG) cDNA library prepared from the crayfish Cherax quadricarinatus, we have identified a novel insulin-like growth factor-binding protein (IGFBP) termed Cq-IGFBP. Based on bioinformatics analyses, the deduced Cq-IGFBP was shown to share high sequence homology with IGFBP family members from both invertebrates and vertebrates. The protein also includes a sequence determinant proven crucial for ligand binding, which according to three-dimensional modeling is assigned to the exposed outer surface of the protein. Recombinant Cq-IGFBP (rCq-IGFBP) protein was produced and, using a "pulldown" methodology, was shown to specifically interact with the insulin-like AG hormone of the crayfish (Cq-IAG). Particularly, using both mass spectral analysis and an immunological tool, rCq-IGFBP was shown to bind the Cq-IAG prohormone. Furthermore, a peptide corresponding to residues 23-38 of the Cq-IAG A-chain was found sufficient for in vitro recognition by rCq-IGFBP. Cq-IGFBP is the first IGFBP family member shown to specifically interact with a gender-specific ILP. Unlike their ILP ligands, IGFBPs are highly conserved across evolution, from ancient arthropods, like crustaceans, to humans. Such conservation places ILP signaling at the center of sex-related phenomena in early animal development.

What the Clock Tells the Eye: Lessons from an Ancient Arthropod

Circadian changes in visual sensitivity have been observed in a wide range of species, vertebrates, and invertebrates, but the processes impacted and the underlying mechanisms largely are unexplored. Among arthropods, effects of circadian signals on vision have been examined in most detail in the lateral compound eye (LE) of the American horseshoe crab, Limulus polyphemus, a chelicerate arthropod. As a consequence of processes influenced by a central circadian clock, Limulus can see at night nearly as well as they do during the day. The effects of the clock on horseshoe crab LE retinas are diverse and include changes in structure, gene expression, and rhabdom biochemistry. An examination of the known effects of circadian rhythms on LEs shows that the effects have three important outcomes: an increase in visual sensitivity at night, a rapid decrease in visual sensitivity at dawn, and maintenance of eyes in a relatively low state of sensitivity during the day, even in the dark. All three outcomes may be critically important for species' survival. Specific effects of circadian rhythms on vision will certainly vary with species and according to life styles. Studies of the circadian regulation of Limulus vision have revealed that these effects can be extremely diverse and profound and suggest that circadian clocks can play a critical role in the ability of animals to adapt to the dramatic daily changes in ambient illumination.

Carotenoids in unexpected places: Gall midges, lateral gene transfer, and carotenoid biosynthesis in animals

Carotenoids are conjugated isoprenoid molecules with many important physiological functions in organisms, including roles in photosynthesis, oxidative stress reduction, vision, diapause, photoperiodism, and immunity. Until recently, it was believed that only plants, microorganisms, and fungi were capable of synthesizing carotenoids and that animals acquired them from their diet, but recent studies have demonstrated that two arthropods (pea aphid and spider mite) possess a pair of genes homologous to those required for the first step of carotenoid biosynthesis. Absent in all other known animal genomes, these genes appear to have been acquired by aphids and spider mites in one or several lateral gene transfer events from a fungal donor. We report the third case of fungal carotenoid biosynthesis gene homologs in an arthropod: flies from the family Cecidomyiidae, commonly known as gall midges. Using phylogenetic analyses we show that it is unlikely that lycopene cyclase/phytoene synthase and phytoene desaturase homologs were transferred singly to an ancient arthropod ancestor; instead we propose that genes were transferred independently from related fungal donors after divergence of the major arthropod lineages. We also examine variation in intron placement and copy number of the carotenoid genes that may underlie function in the midges. This trans-kingdom transfer of carotenoid genes may represent a key innovation, underlying the evolution of phytophagy and plant-galling in gall midges and facilitating their extensive diversification across plant lineages.

A study on the energy source in the developing embryo of the mangrove horseshoe crab, Carcinoscorpius rotundicauda (Latreille)

The mangrove horseshoe crab, Carcinoscorpius rotundicauda, is an Asian species found in Malaysia. This ancient arthropod has a long incubational period during which it depends on various energy sources for both embryogenesis and organogenesis. This study describes the trend of energy utilization from the endogenous reserves by the developing embryos from 0 to 40 days of incubation (until the hatching of the larvae). The dry weight, insoluble protein, carbohydrate, glycogen and lipid showed a declining trend from 0 to 40 days of incubation, whereas the wet weight, water content, ash content and soluble protein showed an increasing trend. Selected micro-elements such as Cu2+, Fe2+ and Zn2+ also demonstrated an interesting trend in the developing eggs when the egg mass was subjected to inductively coupled plasma mass spectrometry analysis, where these elements showed a high correlation with the moulting stages and egg development. Maximum variations within the micro-elements were observed during the 1st (10 days after fertilization) and 2nd (35–36 days after fertilization) moulting stages within the developing eggs. This study clearly indicated that the moulting cycles of C. rotundicauda during embryonic development influence energy utilization in the form of carbohydrates, lipids, proteins, glycogen and other micro-elements.

Antibacterial activity of the venom of Heterometrus xanthopus

Heterometrus xanthopus (Scorpion) is one of the most venomous and ancient arthropods. Its venom contains anti-microbial peptides like hadrurin, scorpine, Pandinin 1, and Pandinin 2 that are able to effectively kill multidrug-resistant pathogens. The present study was conducted to evaluate the anti-bacterial activity of H. xanthopus venom. Six Gram-positive and Gram-negative bacterial strains were tested against 1/100, 1/10, and 1/1 fractions of distilled water diluted and crude venom. 1/100 and 1/10 dilutions were not successful in any of the six bacterial strains studied while the 1/1 dilution was effective on Bacillus subtilis ATCC 6633, Salmonella typhimurium ATCC 14028, and Pseudomonas aeruginosa ATCC 27853 with highest zone of inhibition were obtained on B. subtilis. Crude venom was effective against Enterococcus faecalis ATCC 14506, B. subtilis, S. typhimurium, and P. aeruginosa. The most effective results were observed on B. subtilis.

Modularity of a Cambrian ptychoparioid trilobite cranidium

Modularity of the cranidium of Crassifimbra? metalaspis, a Cambrian ptychoparioid trilobite, is investigated using landmark-based geometric morphometric methods to gain insight into the integration among morphogenetic processes responsible for shaping the head of an ancient arthropod. Of particular interest is the extent to which the structure of phenotypic integration was governed by direct interactions among developmental pathways, because these interactions may generate long-term constraints on evolutionary innovation. A modified two-way ANOVA decomposes cranidial shape variation into components representing symmetric variation among individuals and fluctuating asymmetry (FA). The structure of integration of each of these components is inferred from correlated deviations in shape among nine partitions of the cranidium. Significant correlation among partitions in FA indicates direct interactions among their respective developmental pathways. An a priori hypothesis that modularity was determined by functional association among partitions is not well supported by the among-partition correlation structure for either component of variation. Instead, exploratory analyses reveal that phenotypic integration was strongly influenced by spatially localized morphogenetic controls. Comparison of the structures of the Individuals and FA components of variation reveals that the two share relatively few commonalities: the structure of phenotypic integration was only weakly influenced by direct interactions. The large contribution of parallel variation to phenotypic integration suggests that modularity was unlikely to have imposed a long-term constraint on evolutionary innovation in these early trilobites.

An arthropod cis-regulatory element functioning in sensory organ precursor development dates back to the Cambrian

BackgroundAn increasing number of publications demonstrate conservation of function of cis-regulatory elements without sequence similarity. In invertebrates such functional conservation has only been shown for closely related species. Here we demonstrate the existence of an ancient arthropod regulatory element that functions during the selection of neural precursors. The activity of genes of the achaete-scute (ac-sc) family endows cells with neural potential. An essential, conserved characteristic of proneural genes is their ability to restrict their own activity to single or a small number of progenitor cells from their initially broad domains of expression. This is achieved through a process called lateral inhibition. A regulatory element, the sensory organ precursor enhancer (SOPE), is required for this process. First identified in Drosophila, the SOPE contains discrete binding sites for four regulatory factors. The SOPE of the Drosophila asense gene is situated in the 5' UTR.ResultsThrough a manual comparison of consensus binding site sequences we have been able to identify a SOPE in UTR sequences of asense-like genes in species belonging to all four arthropod groups (Crustacea, Myriapoda, Chelicerata and Insecta). The SOPEs of the spider Cupiennius salei and the insect Tribolium castaneum are shown to be functional in transgenic Drosophila. This would place the origin of this regulatory sequence as far back as the last common ancestor of the Arthropoda, that is, in the Cambrian, 550 million years ago.ConclusionsThe SOPE is not detectable by inter-specific sequence comparison, raising the possibility that other ancient regulatory modules in invertebrates might have escaped detection.

Cretaceous African life captured in amber

Amber is of great paleontological importance because it preserves a diverse array of organisms and associated remains from different habitats in and close to the amber-producing forests. Therefore, the discovery of amber inclusions is important not only for tracing the evolutionary history of lineages with otherwise poor fossil records, but also for elucidating the composition, diversity, and ecology of terrestrial paleoecosystems. Here, we report a unique find of African amber with inclusions, from the Cretaceous of Ethiopia. Ancient arthropods belonging to the ants, wasps, thrips, zorapterans, and spiders are the earliest African records of these ecologically important groups and constitute significant discoveries providing insight into the temporal and geographical origins of these lineages. Together with diverse microscopic inclusions, these findings reveal the interactions of plants, fungi and arthropods during an epoch of major change in terrestrial ecosystems, which was caused by the initial radiation of the angiosperms. Because of its age, paleogeographic location and the exceptional preservation of the inclusions, this fossil resin broadens our understanding of the ecology of Cretaceous woodlands.

Discovery of six families of fungal defensin-like peptides provides insights into origin and evolution of the CSαβ defensins

The defensins with a conserved cysteine-stabilized α-helix and β-sheet (CSαβ) structural motif are a group of unique antimicrobial polypeptides widely distributed in plants and animals. Recently, one defensin-like peptide (DLP) with high degree of sequence and structural similarity to defensins from ancient arthropods and molluscs has been identified in a saprophytic fungus [Mygind, P.H., Fischer, R.L., Schnorr, K.M., Hansen, M.T., Sönksen, C.P., Ludvigsen, S., Raventós, D., Buskov, S., Christensen, B., De Maria, L., Taboureau, O., Yaver, D., Elvig-Jørgensen, S.G., Sørensen, M.V., Christensen, B.E., Kjærulff, S.K., Frimodt-Moller, N., Lehrer, R.I., Zasloff, M., Kristensen, H.-H., 2005. Plectasin is a peptide antibiotic with therapeutic potential from a saprophytic fungus. Nature 437, 975–980], which poses an important question regarding the evolutionary relationships of this class of effectors of innate immunity in three eukaryotic kingdoms. Here, we report the computational identification of six families of fungal DLPs in which three known defensin types (antibacterial ancient invertebrate-type defensins (AITDs), antibacterial classical insect-type defensins (CITDs), and antifungal plant/insect-type defensins (PITDs)) can be clearly assigned. Sharing of these defensin types between animals and fungi supports their closer evolutionary relationship, consistent with the Opisthokonta Hypothesis. Conservation of the PITDs across three eukaryotic kingdoms suggests their earlier origin than the antibacterial defensins, probably preceded plants and Opisthokonta split. Finally, recognition of an early gene duplication event in the Aspergillus terreus genome allows us to establish a paralogous relationship between AITDs and CITDs, which highlights extensive lineage-specific defensin gene loss during evolution.

Conservation and variation in Ubx expression among chelicerates.

Chelicerates are an ancient arthropod group with a distinct body plan composed of an anterior (prosoma) and a posterior portion (opisthosoma). The expression of the Hox gene Ultrabithorax (Ubx) has been examined in a single representative of the chelicerates, the spider Cupiennius salei. In spiders, Ubx expression starts in the second opisthosomal segment (O2). Because the first opisthosomal segment (O1) in spiders is greatly reduced relative to other chelicerates, we hypothesized that the observed Ubx expression pattern might be secondarily modified. Shifts in the anterior boundary of the expression of Ubx have been correlated with functional shifts in morphology within malacostracan crustaceans. Thus, the boundary of Ubx expression between chelicerates with different morphologies in their anterior opisthosoma could also be variable. To test this prediction, we examined the expression patterns of Ubx and abdominal-A (collectively referred to as UbdA) in two basal chelicerate lineages, scorpions and xiphosurans (horseshoe crabs), which exhibit variation in the morphology of their anterior opisthosoma. In the scorpion Paruroctonus mesaensis, the anterior border of early expression of UbdA is in a few cells in the medial, posterior region of the O2 segment, with a predominant expression in O3 and posterior. Expression later spreads to encompass the whole O2 segment and a ventral, posterior portion of the O1 segment. In the xiphosuran Limulus polyphemus, early expression of UbdA has an anterior boundary in the segment. Later in development, the anterior boundary moves forward one segment to the chilarial (O1) segment. Thus, the earliest expression boundary of UbdA lies within the second opisthosomal segment in all the chelicerates examined. These results suggest that rather than being derived, the spider UbdA expression in O2 likely reflects the ancestral expression boundary. Changes in the morphology of the first opisthosomal segment are either not associated with changes in UbdA expression or correlate with late developmental changes in UbdA expression.

Two isoforms of a member of the arthropod defensin family from the soft tick, Ornithodoros moubata (Acari: Argasidae)

We previously purified and determined the partial amino acid sequence of a 4 kDa peptide having high homology with scorpion defensin from the hemolymph of adult fed female soft ticks, Ornithodoros moubata. In this study, the full length sequences of two defensin isoforms were obtained. Deduced amino acid sequences reveal a precursor protein of 73 amino acid residues with a mature portion consisting of 37 amino acid residues. This mature peptide contains six cysteine residues conserved in the same location as other invertebrate defensins. Phylogenetic analysis reveals that Ornithodoros defensin is most closely related to scorpion defensin and other more ancient arthropods. Ornithodoros defensin mRNA is constitutively expressed and up-regulated by blood-feeding and bacterial injection. Ornithodoros defensin gene expression occurs mainly in the midgut. This is the first report of the cloning and gene expression of an antibacterial peptide from the Acari.

Involvement of α 2-macroglobulin and C-reactive protein in a complement-like hemolytic system in the arthropod, Limulus polyphemus

Homologues of two plasma proteins of vertebrates, α 2-macroglobulin and C-reactive protein, participate in a hemolytic system of the ancient arthropod, Limulus polyphemus. C-reactive protein, which can under the appropriate circumstances activate the classical pathway of the mammalian complement system, is an essential element of the hemolytic system of Limulus. The selective removal of C-reactive protein from the plasma with phosphorylethanolamine-agarose inactivated hemolysis. Addition of affinity-purified C-reactive protein to inactive plasma restored activity. Exposure of plasma to phosphorylethanolamine in solution potentiated hemolysis. α 2-Macroglobulin is a member of the same protein family as the complement protein C3 and both require an intact thiol ester for activity. Treatment of Limulus plasma with methylamine under conditions that inactivate thiol-ester-containing proteins reduced the hemolytic activity of some plasma preparations. Addition of purified Limulus α 2-macroglobulin to the methylamine-treated plasma restored hemolytic activity. However, α 2-macroglobulin is not necessary for hemolysis since the hemolytic activity of some pooled plasma preparations was insensitive to methylamine treatment under conditions that inactivated α 2-macroglobulin. Purified C-reactive protein was hemolytic in the absence of α 2-macroglobulin. These observations suggest that the proteins in Limulus plasma that participate in hemolysis represent the components of an ancient invertebrate defense system with distant evolutionarily affinities to the vertebrate complement system.

Fatty acids of the female horseshoe crab Xiphosura (Limulus) polyphemus

The fatty acid composition of 9 different tissues and organs of the female horseshoe crab Xiphosura (Limulus) polyphemus — one of the very few recent representatives of the ancient arthropod class Merostomata — was investigated in reference to the distribution of fatty acids through the marine food web. Fatty acid spectra, in which polyunsaturated fatty acids are predominant, especially eicosapentaenoic acid (20:5 ω3), revealed features characteristic of marine lipids. However, rather large quantities of monoenoic fatty acids also occur in all organs. In the saturated fatty acid fractions, the high content of branched-chain components is worth noticing, particularly in the gills and the carapace (35%); in all probability, the high amount of the branched-chain fatty acids is associated with their protective function in surface lipids. Isoprenoid fatty acids such as pristanic and phytanic acid were absent.