Toxoplasmosis is a disease of prominent health concern that is caused by the protozoan parasite, Toxoplasma gondii. Proliferation of T. gondii is dependent on its ability to invade host cells, which is mediated, in part, by calcium-dependent protein kinase 1 (CDPK1). We have developed ATP competitive inhibitors of TgCDPK1 that block invasion of parasites into host cells, preventing their proliferation. The presence of a unique glycine gatekeeper residue in TgCDPK1 permits selective inhibition of the parasite enzyme over human kinases. These potent TgCDPK1 inhibitors do not inhibit the growth of human cell lines and represent promising candidates as toxoplasmosis therapeutics.
Background and Aims
Pollen-collecting bees are among the most important pollinators globally, but are also the most common pollen thieves and can significantly reduce plant reproduction. The pollination efficiency of pollen collectors depends on the frequency of their visits to female(-phase) flowers, contact with stigmas and deposition of pollen of sufficient quantity and quality to fertilize ovules. Here we investigate the relative importance of these components, and the hypothesis that floral and inflorescence characteristics mediate the pollination role of pollen collection by bees.
For ten Aloe species that differ extensively in floral and inflorescence traits, we experimentally excluded potential bird pollinators to quantify the contributions of insect visitors to pollen removal, pollen deposition and seed production. We measured corolla width and depth to determine nectar accessibility, and the phenology of anther dehiscence and stigma receptivity to quantify herkogamy and dichogamy. Further, we compiled all published bird-exclusion studies of aloes, and compared insect pollination success with floral morphology.
Species varied from exclusively insect pollinated, to exclusively bird pollinated but subject to extensive pollen theft by insects. Nectar inaccessibility and strong dichogamy inhibited pollination by pollen-collecting bees by discouraging visits to female-phase (i.e. pollenless) flowers. For species with large inflorescences of pollen-rich flowers, pollen collectors successfully deposited pollen, but of such low quality (probably self-pollen) that they made almost no contribution to seed set. Indeed, considering all published bird-exclusion studies (17 species in total), insect pollination efficiency varied significantly with floral shape.
Species-specific floral and inflorescence characteristics, especially nectar accessibility and dichogamy, control the efficiency of pollen-collecting bees as pollinators of aloes.
Pollen theft; pollination efficiency; dichogamy; floral morphology; Aloe; Alooideae; Xanthorrhoeaceae; Asphodeloideae
Nectar guides, contrasting patterns on flowers that supposedly direct pollinators towards a concealed nectar reward, are taxonomically widespread. However, there have been few studies of their functional significance and effects on plant fitness. Most previous studies focused on pollinator behaviour and used artificial flowers in laboratory settings. We experimentally investigated the role of putative nectar guides in a natural system: the South African iris Lapeirousia oreogena, whose flowers have a clearly visible pattern of six white arrow-markings pointing towards the narrow entrance of the long corolla tube, and its sole pollinator, a long-proboscid nemestrinid fly. We painted over none, some or all of the white arrow-markings with ink that matched the colour of the corolla background. Although arrow-marking removal had little effect on the approaches by flies to flowers from a distance, it dramatically reduced the likelihood of proboscis insertion. Export of pollen dye analogue (an estimate of male fitness) was reduced to almost zero in flowers from which all nectar guides had been removed, and fruit set (a measure of female fitness) was also significantly reduced. Our results confirm that the markings on L. oreogena flowers serve as nectar guides and suggest that they are under strong selective maintenance through both male and female fitness components in this pollination system.
pollination; nectar guide; pollinator behaviour; long-tongued fly; plant fitness
Levodopa dose and severity of Parkinson’s disease (PD) are recognized risk factors for levodopa-induced dyskinesia (LID) in humans. The purpose of the present study was to evaluate the ability of these variables to predict severity of LID in a rat model of PD. Varied concentrations of 6-hydroxy-dopamine were injected into the midbrain to produce wide ranges of dopamine depletion in striatum. Three weeks later, rats were given daily injections of levodopa (2–10 mg/kg i.p.) plus benserazide (12.5 mg/kg i.p.) for 15 days. Abnormal involuntary movements (AIMs) were measured for limb, axial, orolingual, and rotatory movements. Dose-response analysis for total AIM scores yielded a levodopa ED50 value of 3.2 mg/kg on treatment day 15. There were strong interrelated correlations between individual AIM categories (ρ > 0.7) and for each AIM category in regard to total AIM score (ρ > 0.7). In rats that received levodopa doses that were greater than the ED50, rates of amphetamine-induced rotation were significantly correlated with total AIM scores (ρ = 0.413). However, of those rotating >5 times/min, 34% had relatively low AIM scores (<8). Likewise, there was a significant correlation between percentages of tyrosine hydroxylase (TH) loss and total AIM scores (ρ = 0.388). However, in those rats that had >85% TH loss, 30% had AIM scores <8. Our results show that given an adequate dose and magnitude of striatal dopamine depletion, levodopa produces dyskinesia with a continuous spectrum of severity. Although levodopa dose and level of dopamine depletion are significant risk factors for LID, we conclude that other factors must contribute to LID susceptibility.
Previous whole-cell patch-pipette studies showed that focal electrical stimulation of the subthalamic nucleus (STN) evokes a long-lasting complex EPSC and synaptically-evoked bursts of action potentials in substantia nigra pars reticulata (SNR) neurons. Although synaptically-evoked bursting may play a role in normal physiology, excessive burst firing correlates with symptoms of Parkinson’s disease. We used patch-pipette recordings in rat brain slices to study the effects of baclofen on complex EPSCs and STN-induced burst firing in SNR neurons. Baclofen (1 μM) caused a reversible, 73% reduction in complex EPSCs, and this effect was blocked by the GABAB antagonist CGP35348 (100 μM). Using the loose-patch method to record extracellular potentials, a lower concentration of baclofen (100 nM) inhibited STN-evoked bursts while leaving spontaneous firing of action potentials less affected. We suggest that strategies that selectively inhibit burst firing in the SNR might have therapeutic potential in the treatment of Parkinson’s disease.
subthalamic nucleus; burst firing; complex EPSC; substantia nigra reticulata; brain slice; baclofen; GABAB
The motility and invasion of Plasmodium parasites is believed to require a cytoplasmic actin-myosin motor associated with a cell surface ligand belonging to the TRAP (thrombospondin-related anonymous protein) family. Current models of invasion usually invoke the existence of specific receptors for the TRAP-family ligands on the surface of the host cell; however, the identities of these receptors remain largely unknown. Here, we identify the GPI-linked protein Semaphorin-7A (CD108) as an erythrocyte receptor for the P. falciparum merozoite-specific TRAP homolog (MTRAP) by using a systematic screening approach designed to detect extracellular protein interactions. The specificity of the interaction was demonstrated by showing that binding was saturable and by quantifying the equilibrium and kinetic biophysical binding parameters using surface plasmon resonance. We found that two MTRAP monomers interact via their tandem TSR domains with the Sema domains of a Semaphorin-7A homodimer. Known naturally-occurring polymorphisms in Semaphorin-7A did not quantitatively affect MTRAP binding nor did the presence of glycans on the receptor. Attempts to block the interaction during in vitro erythrocyte invasion assays using recombinant proteins and antibodies showed no significant inhibitory effect, suggesting the inaccessibility of the complex to proteinaceous blocking agents. These findings now provide important experimental evidence to support the model that parasite TRAP-family ligands interact with specific host receptors during cellular invasion.
Apicomplexan parasites are one of the most significant groups of pathogens infecting humans and include Plasmodium falciparum, the parasite responsible for malaria. These parasites critically depend on their human host and must invade our cells to multiply; therefore, understanding this invasion process - with the eventual aim of therapeutically preventing it - has been a focus for scientific investigation. A key component of the invasion machinery is a family of proteins (the “TRAP” family) which traverse the membrane surrounding the parasite: the part remaining within the parasite connects to a molecular motor that powers invasion, whilst the surface-exposed region is thought to interact with proteins on the surface of the target host cell. One major question that remains unanswered is the identity of the host receptors for the TRAPs. In our paper, we use a method specifically designed to detect interactions that occur in the extracellular space between host and pathogen proteins to reveal a host receptor called Semaphorin-7A for the TRAP-family member used by the blood stage of the malarial parasite – a protein called MTRAP. The characterization of this host-parasite interaction may therefore lead to novel therapies based upon preventing parasite invasion.
Diabetes represents a major public health and health system burden. As part of the Alberta's Caring for Diabetes (ABCD) Project, two quality-improvement interventions are being piloted in four Primary Care Networks in Alberta. Gaps between health research, policy and practice have been documented and the need to evaluate the impact of public health interventions in real-world settings to inform decision-making and clinical practice is paramount. In this article, we describe the application of the RE-AIM framework to evaluate the interventions beyond effectiveness.
Methods and analysis
Two quality-improvement interventions were implemented, based on previously proven effective models of care and are directed at improving the physical and mental health of patients with type-2 diabetes. Our goal is to adapt and apply the RE-AIM framework, using a mixed-methods approach, to understand the impact of the interventions to inform policy and clinical decision-making. We present the proposed measures, data sources and data management and analysis strategies used to evaluate the interventions by RE-AIM dimension.
Ethics and dissemination
Ethics approval for the ABCD Project has been granted from the Health Research Ethics Board (HREB #PRO00012663) at the University of Alberta. The RE-AIM framework will be used to structure our dissemination activities by dimension.
It will be presented at relevant conferences and prepared for publication in peer-reviewed journals. Various products, such as presentations, briefing reports and webinars, will be developed to inform key stakeholders of the findings. Presentation of findings by RE-AIM dimension will facilitate discussion regarding the public health impact of the two interventions within the primary care context of Alberta and lessons learned to be used in programme planning and care delivery for patients with type-2 diabetes. It will also promote the application of evaluation models to better assess the impact of community-based primary healthcare interventions through our dissemination activities.
Neisseria meningitis remains a leading cause of sepsis and meningitis, and vaccines are required to prevent infections by this important human pathogen. Factor H binding protein (fHbp) is a key antigen that elicits protective immunity against the meningococcus and recruits the host complement regulator, fH. As the high affinity interaction between fHbp and fH could impair immune responses, we sought to identify non-functional fHbps that could act as effective immunogens. This was achieved by alanine substitution of fHbps from all three variant groups (V1, V2 and V3 fHbp) of the protein; while some residues affected fH binding in each variant group, the distribution of key amino underlying the interaction with fH differed between the V1, V2 and V3 proteins. The atomic structure of V3 fHbp in complex with fH and of the C-terminal barrel of V2 fHbp provide explanations to the differences in the precise nature of their interactions with fH, and the instability of the V2 protein. To develop transgenic models to assess the efficacy of non-functional fHbps, we determined the structural basis of the low level of interaction between fHbp and murine fH; in addition to changes in amino acids in the fHbp binding site, murine fH has a distinct conformation compared with the human protein that would sterically inhibit binding to fHbp. Non-functional V1 fHbps were further characterised by binding and structural studies, and shown in non-transgenic and transgenic mice (expressing chimeric fH that binds fHbp and precisely regulates complement system) to retain their immunogenicity. Our findings provide a catalogue of non-functional fHbps from all variant groups that can be included in new generation meningococcal vaccines, and establish proof-in-principle for clinical studies to compare their efficacy with wild-type fHbps.
Vaccines are currently available against several serogroups of Neisseria meningitidis. However broadly effective serogroup B vaccines are still required as capsule-based approaches cannot be implemented with this serogroup because of the risks of auto-immunity. As a result, vaccines based on proteins in the bacterial outer membrane are being developed. Factor H binding protein (fHbp) is an important meningococcal immunogen which is able to bind the human complement regulator factor H (fH) at high affinity; this interaction could impair the efficacy of fHbp-based vaccines. Here we perform structure:function analyses to define non-functional fHbps and to explain the basis for the host specificity of the fHbp:fH interaction. The vaccine candidacy of non-functional fHbps was compared with wild-type proteins in a relevant transgenic model. These findings should allow the design and evaluation of future fHbp vaccines against this important human pathogen.
Defects in subunits of the conserved oligomeric Golgi (COG) complex represent a growing subset of congenital disorders of glycosylation (CDGs). In addition to altered protein glycosylation and vesicular trafficking, Cog-deficient patient fibroblasts exhibit a striking delay in the Golgi-disrupting effects of brefeldin A (BFA). Despite the diagnostic value of this BFA resistance, the molecular basis of this response is not known. To investigate potential mechanisms of resistance, we analyzed the localization of the large ARF-GEF, GBF1, in several Cog-deficient cell lines. Our results revealed mislocalization of GBF1 to non-Golgi compartments, in particular the ERGIC, within these cells. Biochemical analysis of GBF1 in control and BFA-treated fibroblasts demonstrated that the steady-state level and membrane recruitment is not substantially affected by COG deficiency, supporting a role for the COG complex in the localization but not membrane association of GBF1. We also showed that pretreatment of fibroblasts with bafilomycin resulted in a GBF1-independent BFA resistance that appears additive with the resistance associated with COG deficiency. These data provide new insight into the mechanism of BFA resistance in Cog-deficient cells by suggesting a role for impaired ARF-GEF localization.
Golgi; retrograde traffic; Conserved Oligomeric Golgi complex; GBF1; BFA; Congenital Disorders of Glycosylation
Most of the shell material in snails is composed of calcium carbonate but the organic shell matrix determines the properties of calcium carbonate crystals. It has been shown that the deposition of calcium carbonate is affected by the ingestion of organic compounds. We hypothesize that organic compounds not synthesized by the snails are important for shell strength and must be obtained from the diet. We tested this idea indirectly by evaluating whether the abundance of the organic matter that snails eat is related to the strength of their shells. We measured shell crushing resistance in the snail Mexipyrgus churinceanus and the abundance of the most common aquatic macrophyte, the water lily Nymphaea ampla, in ten bodies of water in the valley of Cuatro Ciénegas, Mexico. We used stable isotopes to test the assumption that these snails feed on water lily organic matter. We also measured other factors that can affect crushing resistance, such as the density of crushing predators, snail density, water pH, and the concentration of calcium and phosphorus in the water. The isotope analysis suggested that snails assimilate water lily organic matter that is metabolized by sediment bacteria. The variable that best explained the variation in crushing resistance found among sites was the local abundance of water lilies. We propose that the local amount of water lily organic matter provides organic compounds important in shell biomineralization, thus determining crushing resistance. Hence, we propose that a third trophic level could be important in the coevolution of snail defensive traits and predatory structures.
To communicate with animals, plants use signals that are distinct from their surroundings. Animals generally learn to use these signals through associative conditioning; however, signals are most effective when they elicit innate behavioural responses. Many plant species have flowers specialized for pollination by ground-dwelling mammals, but the signals used to attract these pollinators have not been elucidated. Here, we demonstrate the chemical basis for attraction of mammal pollinators to flowers of the dioecious parasitic plant Cytinus visseri (Cytinaceae). Two aliphatic ketones dominate the scent of this species; 3-hexanone, which elicits strong innate attraction in rodents, and 1-hexen-3-one, which repels them in isolation, but not in combination with 3-hexanone. The aliphatic ketone-dominated scent of C. visseri contrasts with those of insect-pollinated plants, which are typically dominated by terpenoids, aromatic or non-ketone aliphatic compounds. 3-hexanone is also known from some bat-pollinated species, suggesting independent evolution of plant signals in derived, highly specialized mammal-pollination systems.
Cytinus visseri; Cytinaceae; dioecy; floral syndrome; nectar; pollination
To determine the effect of metformin on the acute metabolic response to submaximal exercise, the effect of exercise on plasma metformin concentrations, and the interaction between metformin and exercise on the subsequent response to a standardized meal.
RESEARCH DESIGN AND METHODS
Ten participants with type 2 diabetes were recruited for this randomized crossover study. Metformin or placebo was given for 28 days, followed by the alternate condition for 28 days. On the last 2 days of each condition, participants were assessed during a nonexercise and a subsequent exercise day. Exercise took place in the morning and involved a total of 35 min performed at three different submaximal intensities.
Metformin increased heart rate and plasma lactate during exercise (both P ≤ 0.01) but lowered respiratory exchange ratio (P = 0.03) without affecting total energy expenditure, which suggests increased fat oxidation. Metformin plasma concentrations were greater at several, but not all, time points on the exercise day compared with the nonexercise day. The glycemic response to a standardized meal was reduced by metformin, but the reduction was attenuated when exercise was added (metformin × exercise interaction, P = 0.05). Glucagon levels were highest in the combined exercise and metformin condition.
This study reveals several ways by which metformin and exercise therapies can affect each other. By increasing heart rate, metformin could lead to the prescription of lower exercise workloads. Furthermore, under the tested conditions, exercise interfered with the glucose-lowering effect of metformin.
While strong and consistent evidence supports the role of lifestyle modification in the prevention and management of type 2 diabetes (T2DM), the best strategies for program implementation to support lifestyle modification within primary care remain to be determined. The objective of the study is to evaluate the implementation of an evidence-based self- management program for patients with T2DM within a newly established primary care network (PCN) environment.
Using a non-randomized design, participants (total N = 110 per group) will be consecutively allocated in bi-monthly blocks to either a 6-month self-management program lead by an Exercise Specialist or to usual care. Our primary outcome is self-reported physical activity and pedometer steps.
The present study will assess whether a diabetes self-management program lead by an Exercise Specialist provided within a newly emerging model of primary care and linked to available community-based resources, can lead to positive changes in self-management behaviours for adults with T2DM. Ultimately, our work will serve as a platform upon which an emerging model of primary care can incorporate effective and efficient chronic disease management practices that are sustainable through partnerships with local community partners.
Clinical Trials Registration
ClinicalTrials.gov identifier: NCT00991380
Type 2 diabetes; Primary care; Physical activity; Diet; Exercise specialist; Health service research
Effective control and eradication of malaria will require new tools to prevent transmission. Current antimalarial therapies targeting the asexual stage of Plasmodium do not prevent transmission of circulating gametocytes from infected humans to mosquitoes. Here, we describe a new class of transmission-blocking compounds, bumped kinase inhibitors (BKIs), which inhibit microgametocyte exflagellation. Oocyst formation and sporozoite production, necessary for transmission to mammals, were inhibited in mosquitoes fed on either BKI-1–treated human blood or mice treated with BKI-1. BKIs are hypothesized to act via inhibition of Plasmodium calcium-dependent protein kinase 4 and predicted to have little activity against mammalian kinases. Our data show that BKIs do not inhibit proliferation of mammalian cell lines and are well tolerated in mice. Used in combination with drugs active against asexual stages of Plasmodium, BKIs could prove an important tool for malaria control and eradication.
Background and Aims
Although pollination of plants that attract flies by resembling their carrion brood and food sites has been reported in several angiosperm families, there has been very little work done on the level of specificity in carrion mimicry systems and the importance of plant cues in mediating such specialization. Specificity may be expected, as carrion-frequenting flies often exploit different niches, which has been interpreted as avoidance of interspecific competition. Interactions between the orchid Satyrium pumilum and a local assemblage of carrion flies were investigated, and the functional significance of floral traits, especially scent, tested. Pollination success and the incidence of pollinator-mediated self-pollination were measured and these were compared with values for orchids with sexual- and food-deceptive pollination systems.
Methods and Key Results
Observations of insect visitation to animal carcasses and to flowers showed that the local assemblage of carrion flies was dominated by blow flies (Calliphoridae), house flies (Muscidae) and flesh flies (Sarcophagidae), but flowers of the orchid were pollinated exclusively by flesh flies, with a strong bias towards females that sometimes deposited live larvae on flowers. A trend towards similar partitioning of fly taxa was found in an experiment that tested the effect of large versus small carrion quantities on fly attraction. GC-MS analysis showed that floral scent is dominated by oligosulfides, 2-heptanone, p-cresol and indole, compounds that also dominate carrion scent. Flesh flies did not distinguish between floral and carrion scent in a choice experiment using olfactory cues only, which also showed that scent alone is responsible for fly attraction. Pollination success was relatively high (31·5 % of flowers), but tracking of stained pollinia also revealed that a relatively high percentage (46 %) of pollen deposited on stigmas originates from the same plant.
Satyrium pumilum selectively attracts flesh flies, probably because its relatively weak scent resembles that of the small carrion on which these flies predominate. In this way, the plants exploit a specific subset of the insect assemblage associated with carrion. Pollination rates and levels of self-pollination were high compared with those in other deceptive orchids and it is therefore unlikely that this mimicry system evolved to promote outcrossing.
Calliphoridae; deception; floral scent; indole; oligosulfides; osmophore; outcrossing; p-cresol; Sarcophagidae; Satyrium pumilum; specialization
A review of published tetartohedrally twinned macromolecular structures is presented, together with details of the recent structure determination of triclinic tetartohedrally twinned crystals of human complement factor I.
Tetartohedral crystal twinning is discussed as a particular case of (pseudo)merohedral twinning when the number of twinned domains is four. Tetartohedrally twinned crystals often possess pseudosymmetry, with the rotational part of the pseudosymmetry operators coinciding with the twinning operators. Tetartohedrally twinned structures from the literature are reviewed and the recent structure determination of tetartohedrally twinned triclinic crystals of human complement factor I is discussed.
The long half-life of botulinum neurotoxin serotype A (BoNT/A) in cells poses a challenge in developing post-exposure therapeutics complementary to existing antitoxin strategies. Delivery vehicles consisting of the toxin heavy chain (HC), including the receptor-binding domain and translocation domain, connected to an inhibitory cargo offer a possible solution for rescuing intoxicated neurons in victims paralyzed from botulism. Here, we report the expression and purification of soluble recombinant prototype green fluorescent protein (GFP) cargo proteins fused to the entire BoNT/A-HC (residues 544–1295) in Escherichia coli with up to a 40 amino acid linker inserted between the cargo and BoNT/A-HC vehicle. We show that these GFP-HC fusion proteins are functionally active and readily taken up by cultured neuronal cells as well as by neuronal cells in mouse motor nerve endings.
delivery vehicle; cargo; neurotoxin; antitoxin
The severe pediatric disorder mucolipidosis II (ML-II; also known as I-cell disease) is caused by defects in mannose 6-phosphate (Man-6-P) biosynthesis. Patients with ML-II exhibit multiple developmental defects, including skeletal, craniofacial and joint abnormalities. To date, the molecular mechanisms that underlie these clinical manifestations are poorly understood. Taking advantage of a zebrafish model of ML-II, we previously showed that the cartilage morphogenesis defects in this model are associated with altered chondrocyte differentiation and excessive deposition of type II collagen, indicating that aspects of development that rely on proper extracellular matrix homeostasis are sensitive to decreases in Man-6-P biosynthesis. To further investigate the molecular bases for the cartilage phenotypes, we analyzed the transcript abundance of several genes in chondrocyte-enriched cell populations isolated from wild-type and ML-II zebrafish embryos. Increased levels of cathepsin and matrix metalloproteinase (MMP) transcripts were noted in ML-II cell populations. This increase in transcript abundance corresponded with elevated and sustained activity of several cathepsins (K, L and S) and MMP-13 during early development. Unlike MMP-13, for which higher levels of protein were detected, the sustained activity of cathepsin K at later stages seemed to result from its abnormal processing and activation. Inhibition of cathepsin K activity by pharmacological or genetic means not only reduced the activity of this enzyme but led to a broad reduction in additional protease activity, significant correction of the cartilage morphogenesis phenotype and reduced type II collagen staining in ML-II embryos. Our findings suggest a central role for excessive cathepsin K activity in the developmental aspects of ML-II cartilage pathogenesis and highlight the utility of the zebrafish system to address the biochemical underpinnings of metabolic disease.
Nucleosomes are the basic packaging units of chromatin, modulating accessibility of regulatory proteins to DNA and thus influencing eukaryotic gene regulation. Elaborate chromatin remodeling mechanisms have evolved that govern nucleosome organization at promoters, regulatory elements, and other functional regions in the genome1. Analyses of chromatin landscape have uncovered a variety of mechanisms, including DNA sequence preferences, that can influence nucleosome positions2–4. To identify major determinants of nucleosome organization in the human genome, we utilized deep sequencing to map nucleosome positions in three primary human cell types and in vitro. A majority of the genome exhibited substantial flexibility of nucleosome positions while a small fraction showed reproducibly positioned nucleosomes. Certain sites that position in vitro can anchor the formation of nucleosomal arrays that have cell type-specific spacing in vivo. Our results unveil an interplay of sequence-based nucleosome preferences and non-nucleosomal factors in determining nucleosome organization within mammalian cells.
Background and Aims
Animal pollination is typically an uncertain process that interacts with self-incompatibility status to determine reproductive success. Seed set is often pollen-limited, but species with late-acting self-incompatibility (SI) may be particularly vulnerable, if self-pollen deposition results in ovule discounting. Pollination is examined and the occurrence of late-acting SI and ovule discounting assessed in Cyrtanthus breviflorus.
The pollination system was characterized by observing floral visitors and assessing nectar production and spectral reflectance of flowers. To assess late-acting SI and ovule discounting, growth of self- and cross-pollen tubes, and seed set following open pollination or hand pollination with varying proportions of self- and cross-pollen, were examined.
Native honeybees Apis mellifera scutellata pollinated flowers as they actively collected pollen. Most flowers (≥70 %) did not contain nectar, while the rest produced minute volumes of dilute nectar. The flowers which are yellow to humans are visually conspicuous to bees with a strong contrast between UV-reflecting tepals and UV-absorbing anthers and pollen. Plants were self-incompatible, but self-rejection was late-acting and both self- and cross-pollen tubes penetrated ovules. Seed set of open-pollinated flowers was pollen-limited, despite pollen deposition exceeding ovule number by 6-fold. Open-pollinated seed set was similar to that of the cross + self-pollen treatment, but was less than that of the cross-pollen-only treatment.
Flowers of C. breviflorus are pollinated primarily by pollen-collecting bees and possess a late-acting SI system, previously unknown in this clade of the Amaryllidaceae. Pollinators of C. breviflorus deposit mixtures of cross- and self-pollen and, because SI is late-acting, self-pollen disables ovules, reducing female fertility. This study thus contributes to growing evidence that seed production in plants with late-acting SI systems is frequently limited by pollen quality, even when pollinators are abundant.
Amarydillaceae; Cyrtanthus breviflorus; honeybee pollination; late-acting self-incompatibility; ovule discounting; pollen limitation; pollen quantity and quality
The radiation of the angiosperms is often attributed to repeated evolutionary shifts between different pollinators, as this process drives diversification of floral forms and can lead to reproductive isolation. Floral scent is an important functional trait in many pollination systems but has seldom been implicated as a key mechanism in pollinator transitions. In this study, we suggest a role for sulphur compounds in mediating a shift between specialized carrion-fly and pompilid-wasp pollination systems in Eucomis (Hyacinthaceae). Flowers of closely related Eucomis species pollinated by carrion flies or pompilid wasps have very similar greenish-white flowers, but differ markedly in floral scent chemistry (determined by GC–MS analysis of headspace extracts). Comparison of the floral colours of the four Eucomis species in the visual systems of flies and wasps suggests that colour plays little role in pollinator discrimination. Nectar properties and morphology also do not differ strongly between fly- and wasp-pollinated flowers. By comparing floral scent bouquets and experimentally manipulating the scent of plants in the field, we demonstrate that shifts between wasp and fly pollination in these four congeners can depend on the production or suppression of sulphur compounds (dimethyl disulphide and dimethyl trisulphide) in the fragrance bouquet. This suggests that mutations affecting the production of particular scent compounds could precipitate shifts between pollinators, independently of floral morphology, colour or nectar properties.
pollinator transition; floral evolution; pollination syndrome; myophily; oligosulphide
The geographic mosaic theory of coevolution predicts 1) spatial variation in predatory structures as well as prey defensive traits, and 2) trait matching in some areas and trait mismatching in others mediated by gene flow. We examined gene flow and documented spatial variation in crushing resistance in the freshwater snails Mexipyrgus churinceanus, Mexithauma quadripaludium, Nymphophilus minckleyi, and its relationship to the relative frequency of the crushing morphotype in the trophically polymorphic fish Herichthys minckleyi. Crushing resistance and the frequency of the crushing morphotype did show spatial variation among 11 naturally replicated communities in the Cuatro Ciénegas valley in Mexico where these species are all endemic. The variation in crushing resistance among populations was not explained by geographic proximity or by genetic similarity in any species. We detected clear phylogeographic patterns and limited gene flow for the snails but not for the fish. Gene flow among snail populations in Cuatro Ciénegas could explain the mosaic of local divergence in shell strength and be preventing the fixation of the crushing morphotype in Herichthys minckleyi. Finally, consistent with trait matching across the mosaic, the frequency of the fish morphotype was negatively correlated with shell crushing resistance likely reflecting the relative disadvantage of the crushing morphotype in communities where the snails exhibit relatively high crushing resistance.
MxiG is a single-pass membrane protein that oligomerizes within the inner membrane ring of the Shigella flexneri type III secretion system (T3SS). The MxiG N-terminal domain (MxiG-N) is the predominant cytoplasmic structure; however, its role in T3SS assembly and secretion is largely uncharacterized. We have determined the solution structure of MxiG-N residues 6–112 (MxiG-N(6–112)), representing the first published structure of this T3SS domain. The structure shows strong structural homology to forkhead-associated (FHA) domains. Canonically, these cell-signaling modules bind phosphothreonine (Thr(P)) via highly conserved residues. However, the putative phosphate-binding pocket of MxiG-N(6–112) does not align with other FHA domain structures or interact with Thr(P). Furthermore, mutagenesis of potential phosphate-binding residues has no effect on S. flexneri T3SS assembly and function. Therefore, MxiG-N has a novel function for an FHA domain. Positioning of MxiG-N(6–112) within the EM density of the S. flexneri needle complex gives insight into the ambiguous stoichiometry of the T3SS, supporting models with 24 MxiG subunits in the inner membrane ring.
Bacteria; NMR; Protein Assembly; Protein Secretion; Signal Transduction; Shigella flexneri; Type Three Secretion System
The structure of rat CrrY1–4 determined in two distinct crystal forms shows a pronounced bend at the interface between domains 3 and 4.
Complement receptor 1-related protein Y (CrrY) is an important cell-surface regulator of complement that is unique to rodent species. The structure of rat CrrY domains 1–4 has been determined in two distinct crystal forms and reveals a 70° bend between domains 3 and 4. Comparisons of this structure with those of other complement regulators suggests that rearrangement of this interface may occur on forming the regulatory complex with C3b.
complement regulator; CrrY; rat; CCP