First chromosome scale genomes of ithomiine butterflies (Nymphalidae: Ithomiini): Comparative models for mimicry genetic studies

The ithomiine butterflies (Nymphalidae: Danainae) represent the largest known radiation of Müllerian mimetic butterflies. They dominate by number the mimetic butterfly communities, which include species such as the iconic neotropical Heliconius genus. Recent studies on the ecology and genetics of speciation in Ithomiini have suggested that sexual pheromones, colour pattern and perhaps hostplant could drive reproductive isolation. However, no reference genome was available for Ithomiini, which has hindered further exploration on the genetic architecture of these candidate traits, and more generally on the genomic patterns of divergence. Here, we generated high-quality, chromosome-scale genome assemblies for two Melinaea species, M. marsaeus and M. menophilus, and a draft genome of the species Ithomia salapia. We obtained genomes with a size ranging from 396 to 503 Mb across the three species and scaffold N50 of 40.5 and 23.2 Mb for the two chromosome-scale assemblies. Using collinearity analyses we identified massive rearrangements between the two closely related Melinaea species. An annotation of transposable elements and gene content was performed, as well as a specialist annotation to target chemosensory genes, which is crucial for host plant detection and mate recognition in mimetic species. A comparative genomic approach revealed independent gene expansions in ithomiines and particularly in gustatory receptor genes. These first three genomes of ithomiine mimetic butterflies constitute a valuable addition and a welcome comparison to existing biological models such as Heliconius, and will enable further understanding of the mechanisms of adaptation in butterflies.     

Of Ducklings and Turing Machines: Interactive Playbacks Enhance Subsequent Responsiveness to Conspecific Calls

Distress calls of mallard ducklings consist of a highly stereotyped series of notes. When two ducklings are simultaneously separated from the brood, they characteristically call in alternation. Each bird inhibits its calls while the other is vocalizing, thus preventing masking and facilitating localization by the hen. I examined whether exposure of ducklings to calls presented in a naturalistic alternating pattern affected subsequent responsiveness to them. Individual ducklings were exposed to either 40 s or 80 s of computer-controlled distress call playbacks, using an algorithm that mimicked the behavior of another duckling. Calls were presented only when the subject was silent and ended as soon as it began to vocalize. Each of these ducklings was paired with a ‘yoked’ control. Birds in these control groups experienced exactly the same pattern of playbacks as the ‘interactive’ birds, but the stimuli had no consistent relationship to their own vocal behavior. When the same calls were played back 24 h later, in a fixed pattern that was independent of the ducklings' behavior, birds that had received 80 s of prior interactive exposure were significantly more responsive than both their yoked controls and birds receiving only 40 s of such playbacks. This result suggests that interactive vocal experience, characteristic of natural communication, affects the subsequent perceptual sensitivity of ducklings.     

The Problematic Paralbunea Hu and Tao, 1996: Homonymy, Generic Nom. Nov., and Correct Taxonomic Placement

The fossil genus Paralbunea Hu and Tao, described as an anomuran albuneid, is a junior homonym of Paralbunea Serène and a possible synonym of either the brachyuran raninid Ranilia H. Milne Edwards, 1837, or Cosmonotus Adams and White, 1848. The type series of P. taipeiensis Hu and Tao is composed of at least two species belonging to two raninid genera and appears (in part) assignable to the genus Ranilia and (in part) to Cosmonotus grayi Adams and White, 1848. However, due to both limited availability and poor quality of the holotype of Paralbunea taipeiensis , and destruction of the paratypes, this taxon cannot be unequivocally synonymized with any particular raninid taxon and a new name is proposed to replace Hu and Tao's preoccupied generic name. Caution is urged when describing purported albuneid fossils, due to strong convergence with raninid crabs, which are far more common in the fossil record.     

Protein stabilization by compatible solutes. Effect of diglycerol phosphate on the dynamics of Desulfovibrio gigas rubredoxin studied by NMR.

Heteronuclear NMR relaxation measurements and hydrogen exchange data have been used to characterize protein dynamics in the presence or absence of stabilizing solutes from hyperthermophiles. Rubredoxin from Desulfovibrio gigas was selected as a model protein and the effect of diglycerol phosphate on its dynamic behaviour was studied. The presence of 100 mM diglycerol phosphate induces a fourfold increase in the half-life for thermal denaturation of D. gigas rubredoxin. A model-free analysis of the protein backbone relaxation parameters shows an average increase of generalized order parameters of 0.015 reflecting a small overall reduction in mobility of fast-scale motions. Hydrogen exchange data acquired over a temperature span of 20 degrees C yielded thermodynamic parameters for the structural opening reactions that allow for the exchange. This shows that the closed form of the protein is stabilized by an additional 1.6 kJ x mol(-1) in the presence of the solute. The results seem to indicate that the stabilizing effect is due mainly to a reduction in mobility of the slower, larger-scale motions within the protein structure with an associated increase in the enthalpy of interactions.     

Molecular characterization of the lysosomal acid phosphatase fromDrosophila melanogaster

InDrosophila, unlike humans, the lysosomal acid phosphatase (Acph-1) is a non-essential enzyme. It is also one of the most rapidly evolving gene-enzyme systems in the genus. In order to determine which parts of the enzyme are conserved and which parts are apparently under little functional constraint, we cloned the gene fromDrosophila melanogaster via a chromosomal walk. Fragments from the gene were used to recover an apparently full-length cDNA. The cDNA was subcloned into aDrosophila transformation vector where it was under the control of the 5′ promoter sequence of thehsp-70 gene. Three independent transformants were obtained; in each, Acph-1 expression from the cDNA was constitutive and not dependent on heat shock, as determined by densitometric analyses of the allozymic forms of the enzyme. The pattern of expression indicates thehsp-70 and endogenousAcph-1 promoters act together in some, but not all, tissues. The sequence of the cDNA was determined using deletions made with exonuclease III, and primers deduced from the cDNA sequence were used to sequence the genomic clone. Five introns were found, and putative 5′ up-stream regulatory sequences were identified. Amino acid sequence comparisons have revealed several highly conserved motifs betweenDrosophila Acph-1 and vertebrate lysosomal and prostatic acid phosphatases.     

Practical applications of time-averaged restrained molecular dynamics to ligand-receptor systems: FK506 bound to the Q50R,A95H,K98I triple mutant of FKBP-13

Summary The ability of time-averaged restrained molecular dynamics (TARMD) to escape local low-energy conformations and explore conformational space is compared with conventional simulated-annealing methods. Practical suggestions are offered for performing TARMD calculations with ligand-receptor systems, and are illustrated for the complex of the immunosuppressant FK506 bound to Q50R,A95H,K98I triple mutant FKBP-13. The structure of ¹³C-labeled FK506 bound to triple-mutant FKBP-13 was determined using a set of 87 NOE distance restraints derived from HSQC-NOESY experiments. TARMD was found to be superior to conventional simulated-annealing methods, and produced structures that were conformationally similar to FK506 bound to wild-type FKBP-12. The individual and combined effects of varying the NOE restraint force constant, using an explicit model for the protein binding pocket, and starting the calculations from different ligand conformations were explored in detail.Abbreviations DG distance geometry - dmFKBP-12 double-mutant (R42K,H87V) FKBP-12 - FKBP-12 FK506-binding protein (12 kDa) - FKBP-13 FK506-binding protein (13 kDa) - HSQC heteronuclear single-quantum coherence - K_NOE force constant (penalty) for NOE-derived distance restraints - MD molecular dynamics - NOE nuclear Overhauser effect - SA simulated annealing - TARMD molecular dynamics with time-averaged restraints - tmFKBP-13 triple-mutant (Q50R,A95H,K98I) FKBP-13 - wtFKBP-12 wild-type FKBP-12     

The elongation of amylose and amylopectin chains in isolated starch granules

The aim of this work was to investigate the conditions required for amylose synthesis in starch granules. Although the major granule-bound isoform of starch synthase - GBSSI - catalyses the synthesis of amylose in vivo, ¹⁴C from ADP[¹⁴C]glucose was incorporated primarily into a specific subset of amylopectin chains when supplied to starch granules isolated from pea (Pisum sativum L.) embryos and potato (Solanum tuberosum L.) tubers. Incubation of granules with soluble extracts of these organs revealed that the extracts contained compounds that increased the incorporation of ¹⁴C into amylose. These compounds were rendered inactive by treatment of the extracts with α-glucosidase, suggesting that they were malto-oligosaccharides. Consistent with this idea, provision of pure malto-oligosaccharides to isolated granules resulted in a dramatic shift in the pattern of incorporation of ¹⁴C, from amylopectin chains to amylose molecules. Comparison of the pattern of incorporation in granules from wild-type peas and lam mutant peas which lack GBSSI showed that this effect of malto-oligosaccharides was specifically on GBSSI. The significance of these results for understanding of the synthesis of amylose and amylopectin in storage organs is discussed.     

Effects of Interactions BetweenPasteurella haemolyticaandBordetella parapertussisonin vitroPhagocytosis by Lung Macrophages

The aim of the work was to determine the effect of exposing ovine bronchoalveolar macrophages (BAM)in vivotoPasteurella haemolyticaand/orBordetella parapertussison the subsequent uptake and killing ofP. haemolyticaby these cellsin vitro. Exposurein vivotoP. haemolyticadid not affect the uptake ofP. haemolyticaby BAMin vitrobut reduced (P< 0·05) the intracellular killing of bacteria. Exposurein vivotoB. parapertussishad no significant effect on either the uptake of killing ofP. haemolytica in vitro. However, sequential exposurein vivotoB. parapertussisandP. haemolyticareduced both the ingestion (P< 0·05) and killing (P< 0·001) ofP. haemolytica in vitro. These results indicate that exposure toP. haemolyticacompromised the bacterial killing mechanisms of BAM and that synergy betweenB. parapertussisandP. haemolyticareduced the ability of BAM to ingest bacteria.     

The mechanism of resistance to paraquat is strongly temperature dependent in resistant Hordeum leporinum Link and H. glaucum Steud.

Paraquat-resistant biotypes of the closely-related weed species Hordeum leporinum Link and H. glaucum Steud. are highly resistant to paraquat when grown during the normal winter growing season. However, when grown and treated with paraquat in summer, these biotypes are markedly less resistant to paraquat. This reduced resistance to paraquat in summer is primarily a result of increased temperature following herbicide treatment. The mechanism governing this decrease in resistance at high temperature was examined in H. leporinum. No differences were observed between susceptible and resistant biotypes in the interaction of paraquat with isolated thylakoids when assayed at 15, 25, or 35 °C. About 98 and 65% of applied paraquat was absorbed through the leaf cuticle of both biotypes at 15 and 30 °C, respectively. Following application to leaves, more herbicide was translocated in a basipetal direction in the susceptible biotype compared to the resistant biotype at 15 °C. However, at 30 °C more paraquat was translocated in a basipetal direction in the resistant biotype. Photosynthetic activity of young leaf tissue from within the leaf sheath which had not been directly exposed to paraquat was measured 24 h after treatment of plants with para. quat. This activity was inhibited in the susceptible biotype when plants were maintained at either 15 °C or 30 °C after treatment. In contrast, photosynthetic activity of such tissue of the resistant biotype was not inhibited when plants were maintained at 15 °C after treatment, but was inhibited at 30 °C. The mechanism of resistance in this biotype of H. leporinum correlates with decreased translocation of paraquat and decreased penetration to the active site. This mechanism is temperature sensitive and breaks down at higher temperatures.We are grateful to Zeneca Agrochemicals, Jealotts Hill, Berkshire, UK who provided [¹⁴C]paraquat. E.P. was supported through a Ph.D. scholarship from the Australian International Development Assistance Bureau and C.P. was the recipient of an Australian Research Council Postdoctoral Fellowship.     

Synthesis and biological evaluation of novel NK-1 tachykinin receptor antagonists: The use of cycloalkyl amino acids as a template

In the course of a program aimed at synthesizing novel, potent NK-1 tachykinin receptor antagonists, we developed upon a bioactive model by comparing the low energy structures of a series of peptide and nonpeptide Substance P antagonists. The comparison was based on the super imposition of the aromatic rings, assuming that the rest of the molecule behaves predominantly as a template to arrange the key aromatic groups in the right spatial position. A series of 2-aminocyclohexane carboxylic acid analogues were then selected as the best templates for reproducing the postulated bioactive structure, leading to several pseudo-peptides with interesting biological activity. According to the molecular modeling, these compounds exhibit a neat parallel facing of the indolyl and naphthyl groups at about 3 Å distance. Ultraviolet absorption and steady state fluorescence measurements support this conclusion, showing a linear correlation between the spectral properties and the binding affinity of these analogues. Stacking of the indole ring with naphthalene gives rise to a complex characterized by a well-defined molar extinction coefficient. Consistently, steady state and lifetime fluorescence measurements suggest that the quenching process is ascribable to ground-state interactions between the chromophores. Implications of the π stacking propensity of aromatic groups in the biological activity of the compounds examined are briefly discussed. © 1995 John Wiley & Sons, Inc.