EFLamide (EFLa) is a neuropeptide known for a long time from crustaceans, chelicerates and myriapods. Recently, EFLa-encoding genes were identified in the genomes of apterygote hexapods including basal insect species. In pterygote insects, however, evidence of EFLa was limited to partial sequences in the bed bug (Cimex), migratory locust and a few phasmid species. Here we present identification of a full length EFLa-encoding transcript in the linden bug, Pyrrhocoris apterus (Heteroptera). We created complete null mutants allowing unambiguous anatomical location of this peptide in the central nervous system. Only 2-3 EFLa-expressing cells are located very close to each other near to the surface of the lateral protocerebrum with dense neuronal arborization. Homozygous null EFLa mutants are fully viable and do not have any visible defect in development, reproduction, lifespan, diapause induction or circadian rhythmicity. Phylogenetic analysis revealed that EFLa-encoding transcripts are produced by alternative splicing of a gene that also produces Prohormone-4. However, this Proh-4/EFLa connection is found only in Hemiptera and Locusta, whereas EFLa-encoding transcripts in apterygote hexapods, chelicerates and crustaceans are clearly distinct from Proh-4 genes. The exact mechanism leading to the fused Proh-4/EFLa transcript is not yet determined, and might be a result of canonical cis-splicing, cis-splicing of adjacent genes (cis-SAG), or trans-splicing.

• MeSH
• Phylogeny MeSH
• Heteroptera genetics   metabolism   MeSH
• Insect Proteins chemistry   genetics   metabolism   MeSH
• Thyrotropin-Releasing Hormone genetics   metabolism   MeSH
• Neuropeptides chemistry   genetics   metabolism   MeSH
• Amino Acid Sequence MeSH
• Sequence Alignment MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Circadian clocks are synchronized with the external environment by light and temperature. The effect of these cues on behavior is well-characterized in Drosophila, however, little is known about synchronization in non-model insect species. Therefore, we explored entrainment of locomotor activity by light and temperature in the linden bug Pyrrhocoris apterus (Heteroptera), an insect species with a strong seasonal response (reproductive diapause), which is triggered by both photoperiod and thermoperiod. Our results show that either light or temperature cycles are strong factors entraining P. apterus locomotor activity. Pyrrhocoris is able to be partially synchronized by cycles with temperature amplitude as small as 3°C and more than 50% of bugs is synchronized by 5°C steps. If conflicting zeitgebers are provided, light is the stronger signal. Linden bugs lack light-sensitive (Drosophila-like) cryptochrome. Notably, a high percentage of bugs is rhythmic even in constant light (LL) at intensity ∼400 lux, a condition which induces 100% arrhythmicity in Drosophila. However, the rhythmicity of bugs is still reduced in LL conditions, whereas rhythmicity remains unaffected in constant dark (DD). Interestingly, a similar phenomenon is observed after temperature cycles entrainment. Bugs released to constant thermophase and DD display weak rhythmicity, whereas strong rhythmicity is observed in bugs released to constant cryophase and DD. Our study describes the daily and circadian behavior of the linden bug as a response to photoperiodic and thermoperiodic entraining cues. Although the molecular mechanism of the circadian clock entrainment in the linden bug is virtually unknown, our study contributes to the knowledge of the insect circadian clock features beyond Drosophila research.

• Publication type
• Journal Article MeSH

The attrition of telomeres, the ends of eukaryote chromosomes, and activity of telomerase, the enzyme that restores telomere length, play a role in the ageing process and act as indicators of biological age. A notable feature of advanced eusocial insects is the longevity of reproductive individuals (queens and kings) compared to those from non-reproductive castes (workers and soldiers) within a given species, with a proposed link towards upregulation of telomerase activity in the somatic tissues of reproductive individuals. Given this, eusocial insects provide excellent model systems for research into ageing. We tested telomerase activity and measured telomere length in Bombus terrestris, which is a primitively eusocial insect species with several distinct features compared to advanced social insects. In somatic tissues, telomerase activity was upregulated only in the fat bodies of pre-diapause queens, and this upregulation was linked to heightened DNA synthesis. Telomere length was shorter in old queens compared to that in younger queens or workers. We speculate that (1) the upregulation of telomerase activity, together with DNA synthesis, is the essential step for intensifying metabolic activity in the fat body to build up a sufficient energy reserve prior to diapause, and that (2) the lifespan differences between B. terrestris workers and queens are related to the long diapause period of the queen. A possible relationship between telomere length regulation and TOR, FOXO, and InR as cell signaling components, was tested.

• MeSH
• DNA biosynthesis   MeSH
• Telomerase metabolism   MeSH
• Fat Body enzymology   MeSH
• Bees enzymology   MeSH
• Telomere Shortening MeSH
• Animals MeSH
• Check Tag
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Post-diapause, overwintered adults of the true bug Pyrrhocoris apterus L. (Heteroptera, Pyrrhocoridae) form conspicuous aggregations at warm spots in early spring. Using a combination of laboratory experiments and field observations, we assessed the influence of this behaviour on the seasonal timing of reproduction. In the laboratory, post-diapause pairs mated after an accumulation of 80° days (dd), and females started to lay eggs after an additional 80 dd (considering 10.3 °C as a lower temperature threshold for post-diapause development). In the field, however, females mated as early as late March (based on data from four seasons), which corresponded to the accumulation of <15 dd (based on summing the temperatures recorded at a meteorology station). Such a 'discrepancy' between laboratory and field data is explained by thermoregulation. Aggregated adults bask in sunny spots, thereby increasing their body temperatures to ~25 °C, which is 7-16 °C above the temperature of the surrounding ground. This thermal excess speeds up their post-diapause development and enables early mating and oviposition. As a result, behavioural thermoregulation hastens the seasonal start of reproduction in P. apterus by > 1 month.

• MeSH
• Behavior, Animal * MeSH
• Heteroptera physiology   MeSH
• Seasons MeSH
• Reproduction physiology   MeSH
• Temperature MeSH
• Body Temperature Regulation * MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Geographicals
• Czech Republic MeSH

Ephemeral habitats can impose challenging conditions for population persistence. Survival strategies in these environments can range from high dispersal capacity to the evolution of dormant stages able to tolerate a harsh environment outside the temporal window of favourable conditions [1]. Annual killifish have evolved to live in seasonal pools on the African savannah and display a range of adaptations to cope with an unpredictable environment [2,3]. For most of the year, killifish populations survive as diapausing embryos buried in dry sediment. When savannah depressions fill with rainwater, the fish hatch, grow rapidly and, after attaining sexual maturity, reproduce daily [2,4]. Nothobranchius furzeri, a model species in ageing research [2,3], is distributed in a region where the climate is particularly dry and rains are unpredictable [5]. Here, we demonstrate that the fast juvenile growth and rapid sexual maturation shown by N. furzeri in captivity is actually an underestimate of their natural developmental rate. We estimated the age of N. furzeri in natural populations by counting daily-deposited increments in the otoliths and performing histological analysis of gonads. We found that N. furzeri are capable of reaching sexual maturity within 14 days after hatching, which to our knowledge is the fastest rate of sexual maturation recorded for a vertebrate. We also demonstrate that N. furzeri can grow from an initial length of 5 mm up to 54 mm over the course of a two-week period. Such rapid juvenile development is likely to be adaptive since some pools were entirely desiccated 3-5 weeks after filling, but retained a viable killifish population that reproduced before the adults succumbed to the disappearance of their pool.

• MeSH
• Cyprinodontiformes growth & development   physiology   MeSH
• Rain MeSH
• Ecosystem * MeSH
• Adaptation, Physiological MeSH
• Sexual Maturation * MeSH
• Reproduction MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Letter MeSH
• Research Support, Non-U.S. Gov't MeSH
• Geographicals
• Mozambique MeSH

Juvenile hormone (JH) produced by the corpus allatum (CA) stimulates vitellogenesis and reduces the synthesis of hexamerin proteins in adult females of Pyrrhocoris apterus. At present it is unknown whether the signaling pathway involving the JH receptor gene Methoprene tolerant (Met) and its binding partner Taiman (Tai), regulates the synthesis of accessory gland proteins (ACPs) and hexamerin proteins or effects male survival. Knockdown of genes by injecting Met dsRNA or Tai dsRNA, reduced the amount of ACPs whilst enhancing the amount of hexamerin mRNA in the fat body and the release of hexamerin proteins into haemolymph, as occurs after the ablation of CA. Lifespan was enhanced by injecting Met but not Tai dsRNA. Diapause associated with the natural absence of JH had a stronger effect on all these parameters than the ablation of CA or the knockdown of genes. This indicates there is an additional regulating agent. Both Met and Tai dsRNA induced a several fold increase in JH (JH III skiped bisepoxide) but a concurrent loss of Met or Tai disabled its function. This supports the view that the Met/Tai complex functions as a JH receptor in the regulation of ACPs and hexamerins.

• MeSH
• Corpora Allata physiology   surgery   MeSH
• Gene Knockdown Techniques MeSH
• Hemolymph chemistry   MeSH
• Heteroptera genetics   physiology   MeSH
• Juvenile Hormones blood   genetics   metabolism   MeSH
• RNA Interference MeSH
• Reproduction MeSH
• Signal Transduction * MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH

Adult reproductive diapause is characterized by lower behavioral activity, ceased reproduction and absence of juvenile hormone (JH). The role of JH receptor Methoprene-tolerant (Met) in female reproduction is well established; however, its function in male reproductive development and behavior is unclear. In the bean bug, Riptortus pedestris, circadian genes are essential for mediating photoperiodically-dependent growth of the male accessory glands (MAGs). The present study explores the role of circadian genes and JH receptor in male diapause in the linden bug, Pyrrhocoris apterus. These data indicate that circadian factors Clock, Cycle and Cry2 are responsible for photoperiod measurement, whereas Met and its partner protein Taiman participate in JH reception. Surprisingly, knockdown of the JH receptor neither lowered locomotor activity nor reduced mating behavior of males. These data suggest existence of a parallel, JH-independent or JH-upstream photoperiodic regulation of reproductive behavior.

• MeSH
• Photoperiod * MeSH
• Heteroptera growth & development   physiology   MeSH
• Juvenile Hormones metabolism   MeSH
• Locomotion MeSH
• Methoprene metabolism   MeSH
• Sexual Behavior, Animal MeSH
• Signal Transduction * MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

BACKGROUND: In models extensively used in studies of aging and extended lifespan, such as C. elegans and Drosophila, adult senescence is regulated by gene networks that are likely to be similar to ones that underlie lifespan extension during dormancy. These include the evolutionarily conserved insulin/IGF, TOR and germ line-signaling pathways. Dormancy, also known as dauer stage in the larval worm or adult diapause in the fly, is triggered by adverse environmental conditions, and results in drastically extended lifespan with negligible senescence. It is furthermore characterized by increased stress resistance and somatic maintenance, developmental arrest and reallocated energy resources. In the fly Drosophila melanogaster adult reproductive diapause is additionally manifested in arrested ovary development, improved immune defense and altered metabolism. However, the molecular mechanisms behind this adaptive lifespan extension are not well understood. RESULTS: A genome wide analysis of transcript changes in diapausing D. melanogaster revealed a differential regulation of more than 4600 genes. Gene ontology (GO) and KEGG pathway analysis reveal that many of these genes are part of signaling pathways that regulate metabolism, stress responses, detoxification, immunity, protein synthesis and processes during aging. More specifically, gene readouts and detailed mapping of the pathways indicate downregulation of insulin-IGF (IIS), target of rapamycin (TOR) and MAP kinase signaling, whereas Toll-dependent immune signaling, Jun-N-terminal kinase (JNK) and Janus kinase/signal transducer and activator of transcription (JAK/STAT) pathways are upregulated during diapause. Furthermore, we detected transcriptional regulation of a large number of genes specifically associated with aging and longevity. CONCLUSIONS: We find that many affected genes and signal pathways are shared between dormancy, aging and lifespan extension, including IIS, TOR, JAK/STAT and JNK. A substantial fraction of the genes affected by diapause have also been found to alter their expression in response to starvation and cold exposure in D. melanogaster, and the pathways overlap those reported in GO analysis of other invertebrates in dormancy or even hibernating mammals. Our study, thus, shows that D. melanogaster is a genetically tractable model for dormancy in other organisms and effects of dormancy on aging and lifespan.

• MeSH
• Longevity genetics   MeSH
• Drosophila melanogaster genetics   physiology   MeSH
• Genome, Insect MeSH
• Gene Ontology MeSH
• Insulin genetics   MeSH
• Gene Expression Regulation * MeSH
• Reproduction genetics   MeSH
• Signal Transduction MeSH
• Aging genetics   physiology   MeSH
• Transcriptome genetics   MeSH
• Germ Cells metabolism   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH

Adult females of Pyrrhocoris apterus, programmed for diapause by short-day (SD) photoperiod and those programmed for reproduction by long-day (LD) retain photoperiodic information in continuous darkness (DD) until death. However, if the interruption of SD by DD is made in the course of diapause programming in adults, then the incidence of diapause depends on the number of SD cycles received before DD, with no evidence that the photoperiodic clock is free-running at DD to complete diapause induction. These results indicate that the photoperiodic clock is stopped after transfer to DD and the information accumulated before transfer to DD is maintained. Diapause programming in the adult stage requires 9-10 SD cycles to induce diapause in 80% of individuals. However, if the diapause programming starts after ecdysis of LD-larvae to the last instar, only 3 SD cycles before transfer to DD are required for diapause in 80% of individuals. Surprisingly, if the newly ecdysed last instar LD-larvae, sensitive to photoperiod, are transferred to DD (thus they did not experience any SD), diapause occurs in 40% of the individuals. Thus, diapause 'information' is present in LD-larvae and is responsible for a lower number of SD required for diapause induction in the larval than in the adult stage.

• MeSH
• Time Factors MeSH
• Diapause, Insect physiology   MeSH
• Photoperiod MeSH
• Heteroptera growth & development   MeSH
• Larva growth & development   MeSH
• Animals MeSH
• Check Tag
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Several biogenic amines, including controversial presence of norepinephrine (NE), were identified by the high performance liquid chromatography equipped with electrospray ionisation mass spectrometry in brain complexes of adult females of Pyrrhocoris apterus. Quantitative analysis was performed by the high performance liquid chromatography coupled to electrochemical detector. Levels of NE, dopamine (DA), octopamine (OA) and 5-hydroxytryptamine (5-HT) in brain complexes were measured in reproductive vs. diapause females. In field collected samples, levels of NE and DA were significantly higher in reproductive (May) than in non-reproductive (Sep, Oct, Feb) females. In laboratory females, NE is higher in long day photoperiod (reproduction) than in short day photoperiod (diapause) already from the first week of the adult age, while DA shows differences between the two contrasting photoperiods only from the second week of the adult age. No association between reproductive status and levels of OA and 5-HT was found.

• MeSH
• Chromatography, Liquid MeSH
• Dopamine metabolism   MeSH
• Heteroptera metabolism   MeSH
• Spectrometry, Mass, Electrospray Ionization methods   MeSH
• Catecholamines metabolism   MeSH
• Brain metabolism   MeSH
• Norepinephrine metabolism   MeSH
• Octopamine metabolism   MeSH
• Seasons MeSH
• Reproduction MeSH
• Tilia parasitology   MeSH
• Animals MeSH
• Check Tag
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH