ABSTRACT
All organisms need to balance processes that consume energy against those that produce energy. With an increase in biological complexity over evolutionary time, regulation of this balance has become much more complex, resulting in specialization of metabolic tasks between organelles, cells, organs, and in the case of eusocial organisms, between the individuals who comprise the 'superorganism.' Exemplifying this, nurse honey bees maintain high abdominal lipids, while foragers have very low lipid stores, likely contributing to efficient performance of their social role, and thus to colony fitness. The proximate mechanisms responsible for these metabolic differences remain poorly understood. Here, we investigated the effects of age, worker class, and dietary macronutrients on the abdominal activity of fatty acid synthase (FAS), the enzyme responsible for de novo synthesis of fatty acids as well as age on lipase activity, enzymes responsible for breakdown of stored lipids. We found that FAS but not lipase activity declines as bees age past peak nursing age. Feeding both nurses and foragers carbohydrates increased FAS activity compared with starved bee, but, whether fed or starved, nurses had much higher FAS activity than similarly treated foragers, implicating reduced lipid synthesis as one component of foragers' low lipid stores. Finally, we used artificial diets with different amounts of protein and fat to precociously induce low, forager-like FAS activity levels in nurse-age bees deprived of protein. We speculate that reduced protein appetite and consumption during the nurse-forager transition is responsible for suppressed lipid synthesis in foragers.
ABSTRACT
In many organisms, interactions among genes lead to multiple functional states, and changes to interactions can lead to transitions into new states. These transitions can be related to bifurcations (or critical points) in dynamical systems theory. Characterizing these collective transitions is a major challenge for systems biology. Here, we develop a statistical method for identifying bistability near a continuous transition directly from high-dimensional gene expression data. We apply the method to data from honey bees, where a known developmental transition occurs between bees performing tasks in the nest and leaving the nest to forage. Our method, which makes use of the expected shape of the distribution of gene expression levels near a transition, successfully identifies the emergence of bistability and links it to genes that are known to be involved in the behavioral transition. This proof of concept demonstrates that going beyond correlative analysis to infer the shape of gene expression distributions might be used more generally to identify collective transitions from gene expression data.
Subject(s)
Bees , Gene Expression , Animals , Bees/genetics , Bees/physiologyABSTRACT
The protein vitellogenin (Vg) plays a central role in lipid transportation in most egg-laying animals. High Vg levels correlate with stress resistance and lifespan potential in honey bees (Apis mellifera). Vg is the primary circulating zinc-carrying protein in honey bees. Zinc is an essential metal ion in numerous biological processes, including the function and structure of many proteins. Measurements of Zn2+ suggest a variable number of ions per Vg molecule in different animal species, but the molecular implications of zinc-binding by this protein are not well-understood. We used inductively coupled plasma mass spectrometry to determine that, on average, each honey bee Vg molecule binds 3 Zn2+ -ions. Our full-length protein structure and sequence analysis revealed seven potential zinc-binding sites. These are located in the ß-barrel and α-helical subdomains of the N-terminal domain, the lipid binding site, and the cysteine-rich C-terminal region of unknown function. Interestingly, two potential zinc-binding sites in the ß-barrel can support a proposed role for this structure in DNA-binding. Overall, our findings suggest that honey bee Vg bind zinc at several functional regions, indicating that Zn2+ -ions are important for many of the activities of this protein. In addition to being potentially relevant for other egg-laying species, these insights provide a platform for studies of metal ions in bee health, which is of global interest due to recent declines in pollinator numbers.
Subject(s)
Insect Proteins , Vitellogenins , Bees , Animals , Vitellogenins/metabolism , Insect Proteins/metabolism , Zinc , Binding Sites , LipidsABSTRACT
Vitellogenin (Vg) is a phylogenetically broad glycolipophosphoprotein. A major function of this protein is holding lipid cargo for storage and transportation. Vg has been extensively studied in honey bees (Apis mellifera) due to additional functions in social traits. Using AlphaFold and EM contour mapping, we recently described the protein structure of honey bee Vg. The full-length protein structure reveals a large hydrophobic lipid binding site and a well-defined fold at the C-terminal region. Now, we outline a shielding mechanism that allows the C-terminal region of Vg to cover a large hydrophobic area exposed in the all-atom model. We propose that this C-terminal movement influences lipid molecules' uptake, transport, and delivery. The mechanism requires elasticity in the Vg lipid core as described for homologous proteins in the large lipid transfer protein (LLTP) superfamily to which Vg belongs. Honey bee Vg has, additionally, several structural arrangements that we interpret as beneficial for the functional flexibility of the C-terminal region. The mechanism proposed here may be relevant for the Vg molecules of many species.
ABSTRACT
Proteins are under selection to maintain central functions and to accommodate needs that arise in ever-changing environments. The positive selection and neutral drift that preserve functions result in a diversity of protein variants. The amount of diversity differs between proteins: multifunctional or disease-related proteins tend to have fewer variants than proteins involved in some aspects of immunity. Our work focuses on the extensively studied protein Vitellogenin (Vg), which in honey bees (Apis mellifera) is multifunctional and highly expressed and plays roles in immunity. Yet, almost nothing is known about the natural variation in the coding sequences of this protein or how amino acid-altering variants might impact structure-function relationships. Here, we map out allelic variation in honey bee Vg using biological samples from 15 countries. The successful barcoded amplicon Nanopore sequencing of 543 bees revealed 121 protein variants, indicating a high level of diversity in Vg. We find that the distribution of non-synonymous single nucleotide polymorphisms (nsSNPs) differs between protein regions with different functions; domains involved in DNA and protein-protein interactions contain fewer nsSNPs than the protein's lipid binding cavities. We outline how the central functions of the protein can be maintained in different variants and how the variation pattern may inform about selection from pathogens and nutrition.
Subject(s)
Vitellogenins , Amino Acid Sequence , Animals , Bees/genetics , Vitellogenins/genetics , Vitellogenins/metabolismABSTRACT
Vitellogenin (Vg) is a conserved protein used by nearly all oviparous animals to produce eggs. It is also pleiotropic and performs functions in oxidative stress resistance, immunity, and, in honey bees, behavioral development of the worker caste. It has remained enigmatic how Vg affects multiple traits. Here, we asked whether Vg enters the nucleus and acts via DNA-binding. We used cell fractionation, immunohistology, and cell culture to show that a structural subunit of honey bee Vg translocates into cell nuclei. We then demonstrated Vg-DNA binding theoretically and empirically with prediction software and chromatin immunoprecipitation with sequencing (ChIP-seq), finding binding sites at genes influencing immunity and behavior. Finally, we investigated the immunological and enzymatic conditions affecting Vg cleavage and nuclear translocation and constructed a 3D structural model. Our data are the first to show Vg in the nucleus and suggest a new fundamental regulatory role for this ubiquitous protein. Supplementary information: The online version contains supplementary material available at 10.1007/s13592-022-00914-9.
ABSTRACT
Vitellogenin (Vg) has been implicated as a central protein in the immunity of egg-laying animals. Studies on a diverse set of species suggest that Vg supports health and longevity through binding to pathogens. Specific studies of honey bees (Apis mellifera) further indicate that the vitellogenin (vg) gene undergoes selection driven by local pathogen pressures. Determining the complete 3D structure of full-length Vg (flVg) protein will provide insights regarding the structure-function relationships underlying allelic variation. Honey bee Vg has been described in terms of function, and two subdomains have been structurally described, while information about the other domains is lacking. Here, we present a structure prediction, restrained by experimental data, of flVg from honey bees. To achieve this, we performed homology modeling and used AlphaFold before using a negative-stain electron microscopy map to restrict, orient, and validate our 3D model. Our approach identified a highly conserved Ca2+ -ion-binding site in a von Willebrand factor domain that might be central to Vg function. Thereafter, we used rigid-body fitting to predict the relative position of high-resolution domains in a flVg model. This mapping represents the first experimentally validated full-length protein model of a Vg protein and is thus relevant for understanding Vg in numerous species. Our results are also specifically relevant to honey bee health, which is a topic of global concern due to rapidly declining pollinator numbers.
Subject(s)
Insect Proteins , Vitellogenins , Animals , Bees , Insect Proteins/genetics , Insect Proteins/metabolism , Insecta/metabolism , Longevity , Vitellogenins/genetics , Vitellogenins/metabolismABSTRACT
Various bioactive food compounds may confer health and longevity benefits, possibly through altering or preserving the epigenome. While bioactive food compounds are widely being marketed for human consumption as 'improving health and longevity' by counteracting harmful effects of poor nutrition and lifestyle, claimed effects are often not adequately documented. Using the honey bee (Apis mellifera) as a model species, we here employed a multi-step screening approach to investigate seven compounds for effects on lifespan and DNA methylation using ELISA and whole genome bisulfite sequencing (WGBS). A positive longevity effect was detected for valproic acid, isovaleric acid, and cyanocobalamin. For curcumin, we found that lifespan shortening caused by ethanol intake, was restored when curcumin and ethanol were co-administered. Furthermore, we identified region specific DNA methylation changes as a result of ethanol intake. Ethanol specific changes in DNA methylation were fully or partially blocked in honey bees receiving ethanol and curcumin together. Ethanol-affected and curcumin-blocked differentially methylated regions covered genes involved in fertility, temperature regulation and tubulin transport. Our results demonstrate fundamental negative effects of low dose ethanol consumption on lifespan and associated DNA methylation changes and present a proof-of-principle on how longevity and DNA methylation changes can be negated by the bioactive food component curcumin. Our findings provide a fundament for further studies of curcumin in invertebrates.
Subject(s)
Alcohol Drinking/adverse effects , Curcumin/administration & dosage , Food Ingredients , Longevity/drug effects , Animals , Bees , DNA Methylation/drug effects , Disease Models, Animal , Ethanol/toxicity , Humans , Proof of Concept StudyABSTRACT
Social immunity is a suite of behavioral and physiological traits that allow colony members to protect one another from pathogens, and includes the oral transfer of immunological compounds between nestmates. In honey bees, royal jelly is a glandular secretion produced by a subset of workers that is fed to the queen and young larvae, and which contains many antimicrobial compounds. A related form of social immunity, transgenerational immune priming (TGIP), allows queens to transfer pathogen fragments into their developing eggs, where they are recognized by the embryo's immune system and induce higher pathogen resistance in the new offspring. These pathogen fragments are transported by vitellogenin (Vg), an egg-yolk precursor protein that is also used by nurses to synthesize royal jelly. Therefore, royal jelly may serve as a vehicle to transport pathogen fragments from workers to other nestmates. To investigate this, we recently showed that ingested bacteria are transported to nurses' jelly-producing glands, and here, we show that pathogen fragments are incorporated into the royal jelly. Moreover, we show that consuming pathogen cells induces higher levels of an antimicrobial peptide found in royal jelly, defensin-1.
Subject(s)
Fatty Acids , Vitellogenins , Animals , Bacteria , Bees , LarvaABSTRACT
Human life expectancy increases, but the disease-free part of lifespan (healthspan) and the quality of life in old people may not show the same development. The situation poses considerable challenges to healthcare systems and economies, and calls for new strategies to increase healthspan and for sustainable future approaches to elder care. This call has motivated innovative research on the role of social relationships during ageing. Correlative data from clinical surveys indicate that social contact promotes healthy ageing, and it is time to reveal the causal mechanisms through experimental research. The fruit fly Drosophila melanogaster is a prolific model animal, but insects with more developed social behaviour can be equally instrumental for this research. Here, we discuss the role of social contact in ageing, and identify lines of study where diverse insect models can help uncover the mechanisms that are involved. This article is part of the theme issue 'Ageing and sociality: why, when and how does sociality change ageing patterns?'
Subject(s)
Aging , Insecta/physiology , Animals , Models, Animal , Social BehaviorABSTRACT
Social immunity is a suite of behavioral and physiological traits that allow colony members to protect one another from pathogens, and includes the oral transfer of immunological compounds between nestmates. In honey bees, royal jelly is a glandular secretion produced by a subset of workers that is fed to the queen and young larvae, and which contains many antimicrobial compounds. A related form of social immunity, transgenerational immune priming (TGIP), allows queens to transfer pathogen fragments into their developing eggs, where they are recognized by the embryo's immune system and induce higher pathogen resistance in the new offspring. These pathogen fragments are transported by vitellogenin (Vg), an egg-yolk precursor protein that is also used by nurses to synthesize royal jelly. Therefore, royal jelly may serve as a vehicle to transport pathogen fragments from workers to other nestmates. To investigate this, we recently showed that ingested bacteria are transported to nurses' jelly-producing glands, and here, we show that pathogen fragments are incorporated into the royal jelly. Moreover, we show that consuming pathogen cells induces higher levels of an antimicrobial peptide found in royal jelly, defensin-1.
Subject(s)
Fatty Acids , Vitellogenins , Animals , Bacteria , Bees , LarvaABSTRACT
Honey bees (Apis mellifera) provide an excellent model for studying how complex social behavior evolves and is regulated. Social behavioral traits such as the division of labor have been mapped to specific genomic regions in quantitative trait locus (QTL) studies. However, relating genomic mapping to gene function and regulatory mechanism remains a big challenge for geneticists. In honey bee workers, division of labor is known to be regulated by reproductive physiology, but the genetic basis of this regulation remains unknown. In this case, QTL studies have identified tyramine receptor 1 (TYR1) as a candidate gene in region pln2, which is associated with multiple worker social traits and reproductive anatomy. Tyramine (TA), a neurotransmitter, regulates physiology and behavior in diverse insect species including honey bees. Here, we examine directly the effects of TYR1 and TA on worker reproductive physiology, including ovariole number, ovary function and the production of vitellogenin (VG, an egg yolk precursor). First, we used a pharmacology approach to demonstrate that TA affects ovariole number during worker larval development and increases ovary maturation during the adult stage. Second, we used a gene knockdown approach to show that TYR1 regulates vg transcription in adult workers. Finally, we estimated correlations in gene expression and propose that TYR1 may regulate vg transcription by coordinating hormonal and nutritional signals. Taken together, our results suggest TYR1 and TA play important roles in regulating worker reproductive physiology, which in turn regulates social behavior. Our study exemplifies a successful forward-genetic strategy going from QTL mapping to gene function.
Subject(s)
Bees , Receptors, Biogenic Amine/genetics , Reproduction/genetics , Social Behavior , Tyramine , Animals , Bees/genetics , Bees/metabolism , Behavior, Animal/physiology , Fat Body/drug effects , Fat Body/metabolism , Female , Gene Expression , Genes, Insect , Larva/genetics , Larva/metabolism , Neurotransmitter Agents/metabolism , Neurotransmitter Agents/pharmacology , Ovary/anatomy & histology , Ovary/drug effects , Ovary/metabolism , Quantitative Trait Loci , RNA Interference , Receptors, Biogenic Amine/metabolism , Tyramine/metabolism , Tyramine/pharmacology , Vitellogenins/bloodABSTRACT
In animals, dietary restriction or suppression of genes involved in nutrient sensing tends to increase lifespan. In contrast, food restriction in honeybees (Apis mellifera) shortens lifespan by accelerating a behavioural maturation program that culminates in leaving the nest as a forager. Foraging is metabolically demanding and risky, and foragers experience increased rates of aging and mortality. Food-deprived worker bees forage at younger ages and are expected to live shorter lives. We tested whether suppression of a molecular nutrient sensing pathway is sufficient to accelerate the behavioural transition to foraging and shorten worker life. To achieve this, we reduced expression of the insulin receptor substrate (irs) gene via RNA interference in two selected lines of honeybees used to control for behavioural and genetic variation. irs encodes a membrane-associated protein in the insulin/insulin-like signalling (IIS) pathway that is central to nutrient sensing in animals. We measured foraging onset and lifespan and found that suppression of irs reduced worker bee lifespan in both genotypes, and that this effect was largely driven by an earlier onset of foraging behaviour in a genotype-conditional manner. Our results provide the first direct evidence that an IIS pathway gene influences behavioural maturation and lifespan in honeybees and highlight the importance of considering social environments and behaviours when investigating the regulation of aging and lifespan in social animals.
ABSTRACT
There is a growing need to understand relationships between agricultural intensification and global change. Monitoring solutions, however, often do not include pollinator communities that are of importance to ecosystem integrity. Here, we put forth the honey bee as an economical and broadly available bioindicator that can be used to assess and track changes in the quality of agricultural ecosystems. We detail a variety of simple, low-cost procedures that can be deployed within honey bee hives to gain generalizable information about ecosystem quality at multiple scales, and discuss the potential of the honey bee system in both environmental and ecological bioindication.
Subject(s)
Agriculture , Bees , Environmental Biomarkers , Animals , Climate Change , Ecosystem , Environmental Pollution/adverse effects , Environmental Pollution/analysisABSTRACT
Iron granules containing superparamagnetic magnetite act as magnetoreceptor for magnetoreception in honey bees. Biomineralization of iron granules occurs in the iron deposition vesicles of trophocytes and requires the participation of actin, myosin, ferritin2, and ATP synthase. The mechanism of magnetoreception in honey bees can be explored by suppressing the formation of iron granules. Toward this goal, we injected double-stranded RNA of ferritin2 and ferritin1 into newly emerged worker honey bees to knock down these genes via RNA interference. We confirmed that mRNA and protein production of the ferritins was inhibited, leading to immature iron granules. Downregulating ferritin2 and ferritin1, moreover, leads to different deposition morphology of 7.5-nm diameter iron particles, indicating that the two genes play different roles in the formation of iron granules in worker honey bees.
Subject(s)
Adipocytes/metabolism , Bees/physiology , Behavior, Animal/physiology , Ferritins/genetics , Ferritins/metabolism , Iron/metabolism , RNA Interference , Animals , Ferrosoferric Oxide/metabolism , Gene Knockdown Techniques , Green Fluorescent Proteins/genetics , RNA, Double-Stranded/administration & dosage , RNA, Double-Stranded/genetics , RNA, Messenger/genetics , Real-Time Polymerase Chain ReactionABSTRACT
Insects have rapidly changing energy demands, so they primarily rely on hemolymph and other carbohydrates to carry out life activities. However, how gustatory responsiveness and hemolymph sugar levels coordinate with one another to maintain energetic homeostasis in insects remains largely unknown for the highly social honeybee that goes through large physiological and behavioral changes. The potential role of biogenic amines and neuropeptides in the connection between the regulation of appetite and fluctuating sugar levels in the hemolymph, due to starvation, as the bee ages, was investigated. The largest appetite increase due to the starvation treatment was within the forager age class and this corresponded with an increase in octopamine levels in the brain along with a decline in hemolymph sugar levels. Adipokinetic hormone (AKH) was found in very small quantities in the brain and there were no significant changes in response to starvation treatment. Our findings suggest that the particularly dynamic levels of hemolymph sugar levels may serve as a monitor of the forager honeybee energetic state. Therefore, there may be a pathway in forager bees via octopamine responsible for their precise precipitous regulation of appetite, but to determine cause and effect relationships further investigation is needed.
Subject(s)
Appetite/physiology , Bees/physiology , Brain/metabolism , Hemolymph/metabolism , Octopamine/metabolism , Animals , Hemolymph/chemistry , Sugars/metabolismABSTRACT
Foraging exposes organisms to rewarding and aversive events, providing a selective advantage for maximizing the former while minimizing the latter. Honey bees (Apis mellifera) associate environmental stimuli with appetitive or aversive experiences, forming preferences for scents, locations, and visual cues. Preference formation is influenced by inter-individual variation in sensitivity to rewarding and aversive stimuli, which can be modulated by pharmacological manipulation of biogenic amines. We propose that foraging experiences act on biogenic amine pathways to induce enduring changes to stimulus responsiveness. To simulate varied foraging conditions, freely-moving bees were housed in cages where feeders offered combinations of sucrose solution, floral scents, and aversive electric shock. Transient effects were excluded by providing bees with neutral conditions for three days prior to all subsequent assays. Sucrose responsiveness was reduced in bees that had foraged for scented rather than unscented sucrose under benign conditions. This was not the case under aversive foraging conditions, suggesting an adaptive tuning process which maximizes preference for high quality, non-aversive floral sites. Foraging conditions also influenced antennal lobe octopamine and serotonin, neuromodulators involved in stimulus responsiveness and foraging site evaluation. Our results suggest that individuals' foraging experiences durably modify neurochemistry and shape future foraging behaviour.
Subject(s)
Bees/physiology , Biogenic Amines/metabolism , Feeding Behavior/physiology , Mushroom Bodies/metabolism , Neuropil/metabolism , Sucrose/administration & dosage , Animals , Appetitive Behavior/drug effects , Appetitive Behavior/physiology , Arthropod Antennae/drug effects , Arthropod Antennae/physiology , Octopamine/metabolism , Odorants , RewardABSTRACT
Heavy metal toxicity is an ecological concern in regions affected by processes like mining, industry, and agriculture. At sufficiently high concentrations, heavy metals are lethal to honey bees, but little is known about how sublethal doses affect honey bees or whether they will consume contaminated food. We investigated whether honey bees reject sucrose solutions contaminated with three heavy metals - cadmium, copper, and lead - as a measure of their ability to detect the metals, and whether ingesting these metals altered the bees' sucrose sensitivity. The metals elicited three different response profiles in honey bees. Cadmium was not rejected in any of the assays, and ingesting cadmium did not alter sucrose sensitivity. Copper was rejected following antennal stimulation, but was readily consumed following proboscis stimulation. Ingestion of copper did not alter sucrose sensitivity. Lead appeared to be palatable at some concentrations and altered the bees' sensitivity to and/or valuation of sucrose following antennal stimulation or ingestion of the metal. These differences likely represent unique mechanisms for detecting each metal and the pathology of toxicity. The bees' ability to detect and consume these toxic metals highlights the risk of exposure to these elements for bees living in or near contaminated environments.
Subject(s)
Bees/drug effects , Environmental Pollutants/toxicity , Feeding Behavior/drug effects , Heavy Metal Poisoning/veterinary , Metals, Heavy/toxicity , Animals , Arthropod Antennae/drug effects , Arthropod Antennae/physiology , Bees/physiology , Dose-Response Relationship, Drug , Feeding Behavior/physiology , Heavy Metal Poisoning/physiopathology , Soil/chemistry , Taste/physiology , United States , Water/chemistryABSTRACT
Female insects that survive a pathogen attack can produce more pathogen-resistant offspring in a process called trans-generational immune priming. In the honey bee (Apis mellifera), the egg-yolk precursor protein Vitellogenin transports fragments of pathogen cells into the egg, thereby setting the stage for a recruitment of immunological defenses prior to hatching. Honey bees live in complex societies where reproduction and communal tasks are divided between a queen and her sterile female workers. Worker bees metabolize Vitellogenin to synthesize royal jelly, a protein-rich glandular secretion fed to the queen and young larvae. We ask if workers can participate in trans-generational immune priming by transferring pathogen fragments to the queen or larvae via royal jelly. As a first step toward answering this question, we tested whether worker-ingested bacterial fragments can be transported to jelly-producing glands, and what role Vitellogenin plays in this transport. To do this, we fed fluorescently labelled Escherichia coli to workers with experimentally manipulated levels of Vitellogenin. We found that bacterial fragments were transported to the glands of control workers, while they were not detected at the glands of workers subjected to RNA interference-mediated Vitellogenin gene knockdown, suggesting that Vitellogenin plays a role in this transport. Our results provide initial evidence that trans-generational immune priming may operate at a colony-wide level in honey bees.
Subject(s)
Bees/immunology , Vitellogenins/metabolism , Animals , Bees/metabolism , FemaleABSTRACT
Eusocial insects divide their labour so that individuals working inside the nest are affected by external conditions through a cascade of social interactions. Honey bees (Apis mellifera) transfer food and information via mouth-to-mouth social feeding, ie trophallaxis, a process known to be modulated by the rate of food flow at feeders and familiarity of food's scent. Little is understood about how aversive foraging conditions such as predation and con-specific competition affect trophallaxis. We hypothesized that aversive conditions have an impact on food transfer inside the colony. Here we explore the effect of foragers' aversive experience on downstream trophallaxis in a cage paradigm. Each cage contained one group of bees that was separated from feeders by mesh and allowed to feed only through trophallaxis, and another group that had access to feeders and self-specialized to either forage or distribute food. Our results show that aversive foraging conditions increase non-foragers' trophallaxis with bees restricted from feeder access when food is scented, and have the opposite effect when food is unscented. We discuss potential behavioural mechanisms and implications for the impact of aversive conditions such as malaise inducing toxins, predation, and con-specific competition.