Hunting mode and habitat selection mediate the success of human hunters.

BACKGROUND: As a globally widespread apex predator, humans have unprecedented lethal and non-lethal effects on prey populations and ecosystems. Yet compared to non-human predators, little is known about the movement ecology of human hunters, including how hunting behavior interacts with the environment. METHODS: We characterized the hunting modes, habitat selection, and harvest success of 483 rifle hunters in California using high-resolution GPS data. We used Hidden Markov Models to characterize fine-scale movement behavior, and k-means clustering to group hunters by hunting mode, on the basis of their time spent in each behavioral state. Finally, we used Resource Selection Functions to quantify patterns of habitat selection for successful and unsuccessful hunters of each hunting mode. RESULTS: Hunters exhibited three distinct and successful hunting modes ("coursing", "stalking", and "sit-and-wait"), with coursings as the most successful strategy. Across hunting modes, there was variation in patterns of selection for roads, topography, and habitat cover, with differences in habitat use of successful and unsuccessful hunters across modes. CONCLUSIONS: Our study indicates that hunters can successfully employ a diversity of harvest strategies, and that hunting success is mediated by the interacting effects of hunting mode and landscape features. Such results highlight the breadth of human hunting modes, even within a single hunting technique, and lend insight into the varied ways that humans exert predation pressure on wildlife.

Increasing amyloplast size in wheat endosperm through mutation of PARC6 affects starch granule morphology.

The determination of starch granule morphology in plants is poorly understood. The amyloplasts of wheat endosperm contain large discoid A-type granules and small spherical B-type granules. To study the influence of amyloplast structure on these distinct morphological types, we isolated a mutant in durum wheat (Triticum turgidum) defective in the plastid division protein PARC6, which had giant plastids in both leaves and endosperm. Endosperm amyloplasts of the mutant contained more A- and B-type granules than those of the wild-type. The mutant had increased A- and B-type granule size in mature grains, and its A-type granules had a highly aberrant, lobed surface. This morphological defect was already evident at early stages of grain development and occurred without alterations in polymer structure and composition. Plant growth and grain size, number and starch content were not affected in the mutants despite the large plastid size. Interestingly, mutation of the PARC6 paralog, ARC6, did not increase plastid or starch granule size. We suggest TtPARC6 can complement disrupted TtARC6 function by interacting with PDV2, the outer plastid envelope protein that typically interacts with ARC6 to promote plastid division. We therefore reveal an important role of amyloplast structure in starch granule morphogenesis in wheat.

Loss of PROTEIN TARGETING TO STARCH 2 has variable effects on starch synthesis across organs and species.

Recent work has identified several proteins involved in starch granule initiation, the first step of starch synthesis. However, the degree of conservation in the granule initiation process remains poorly understood, especially among grass species differing in patterns of carbohydrate turnover in leaves, and granule morphology in the endosperm. We therefore compared mutant phenotypes of Hordeum vulgare (barley), Triticum turgidum (durum wheat), and Brachypodium distachyon defective in PROTEIN TARGETING TO STARCH 2 (PTST2), a key granule initiation protein. We report striking differences across species and organs. Loss of PTST2 from leaves resulted in fewer, larger starch granules per chloroplast and normal starch content in wheat, fewer granules per chloroplast and lower starch content in barley, and almost complete loss of starch in Brachypodium. The loss of starch in Brachypodium leaves was accompanied by high levels of ADP-glucose and detrimental effects on growth and physiology. Additionally, we found that loss of PTST2 increased granule initiation in Brachypodium amyloplasts, resulting in abnormal compound granule formation throughout the seed. These findings suggest that the importance of PTST2 varies greatly with the genetic and developmental background and inform the extent to which the gene can be targeted to improve starch in crops.

Rising rates of starch degradation during daytime and trehalose 6-phosphate optimize carbon availability.

Many plants, including Arabidopsis (Arabidopsis thaliana), accumulate starch in the light and remobilize it to support maintenance and growth at night. Starch synthesis and degradation are usually viewed as temporally separate processes. Recently, we reported that starch is also degraded in the light. Degradation rates are generally low early in the day but rise with time. Here, we show that the rate of degradation in the light depends on time relative to dawn rather than dusk. We also show that degradation in the light is inhibited by trehalose 6-phosphate, a signal for sucrose availability. The observed responses of degradation in the light can be simulated by a skeletal model in which the rate of degradation is a function of starch content divided by time remaining until dawn. The fit is improved by extension to include feedback inhibition of starch degradation by trehalose 6-phosphate. We also investigate possible functions of simultaneous starch synthesis and degradation in the light, using empirically parameterized models and experimental approaches. The idea that this cycle buffers growth against falling rates of photosynthesis at twilight is supported by data showing that rates of protein and cell wall synthesis remain high during a simulated dusk twilight. Degradation of starch in the light may also counter over-accumulation of starch in long photoperiods and stabilize signaling around dusk. We conclude that starch degradation in the light is regulated by mechanisms similar to those that operate at night and is important for stabilizing carbon availability and signaling, thus optimizing growth in natural light conditions.

Sucrose synthases are not involved in starch synthesis in Arabidopsis leaves.

Many plants accumulate transitory starch reserves in their leaves during the day to buffer their carbohydrate supply against fluctuating light conditions, and to provide carbon and energy for survival at night. It is universally accepted that transitory starch is synthesized from ADP-glucose (ADPG) in the chloroplasts. However, the consensus that ADPG is made in the chloroplasts by ADPG pyrophosphorylase has been challenged by a controversial proposal that ADPG is made primarily in the cytosol, probably by sucrose synthase (SUS), and then imported into the chloroplasts. To resolve this long-standing controversy, we critically re-examined the experimental evidence that appears to conflict with the consensus pathway. We show that when precautions are taken to avoid artefactual changes during leaf sampling, Arabidopsis thaliana mutants that lack SUS activity in mesophyll cells (quadruple sus1234) or have no SUS activity (sextuple sus123456) have wild-type levels of ADPG and starch, while ADPG is 20 times lower in the pgm and adg1 mutants that are blocked in the consensus chloroplastic pathway of starch synthesis. We conclude that the ADPG needed for starch synthesis in leaves is synthesized primarily by ADPG pyrophosphorylase in the chloroplasts.

A dominant mutation in ß-AMYLASE1 disrupts nighttime control of starch degradation in Arabidopsis leaves.

Arabidopsis (Arabidopsis thaliana) leaves possess a mechanism that couples the rate of nighttime starch degradation to the anticipated time of dawn, thus preventing premature exhaustion of starch and nighttime starvation. To shed light on the mechanism, we screened a mutagenized population of a starvation reporter line and isolated a mutant that starved prior to dawn. The mutant had accelerated starch degradation, and the rate was not adjusted to time of dawn. The mutation responsible led to a single amino acid change (S132N) in the starch degradation enzyme BETA-AMYLASE1 (BAM1; mutant allele named bam1-2D), resulting in a dominant, gain-of-function phenotype. Complete loss of BAM1 (in bam1-1) did not affect rates of starch degradation, while expression of BAM1(S132N) in bam1-1 recapitulated the accelerated starch degradation phenotype of bam1-2D. In vitro analysis of recombinant BAM1 and BAM1(S132N) proteins revealed no differences in kinetic or stability properties, but in leaf extracts, BAM1(S132N) apparently had a higher affinity than BAM1 for an established binding partner required for normal rates of starch degradation, LIKE SEX FOUR1 (LSF1). Genetic approaches showed that BAM1(S132N) itself is likely responsible for accelerated starch degradation in bam1-2D and that this activity requires LSF1. Analysis of plants expressing BAM1 with alanine or aspartate rather than serine at position 132 indicated that the gain-of-function phenotype is not related to phosphorylation status at this position. Our results strengthen the view that control of starch degradation in wild-type plants involves dynamic physical interactions of degradative enzymes and related proteins with a central role for complexes containing LSF1.

Introduction of glucan synthase into the cytosol in wheat endosperm causes massive maltose accumulation and represses starch synthesis.

We expressed a bacterial glucan synthase (Agrobacterium GlgA) in the cytosol of developing endosperm cells in wheat grains, to discover whether it could generate a glucan from cytosolic ADP-glucose. Transgenic lines had high glucan synthase activity during grain filling, but did not accumulate glucan. Instead, grains accumulated very high concentrations of maltose. They had large volumes during development due to high water content, and very shrivelled grains at maturity. Starch synthesis was severely reduced. We propose that cytosolic glucan synthesized by the glucan synthase was immediately hydrolysed to maltose by cytosolic ß-amylase(s). Maltose accumulation resulted in a high osmotic potential in developing grain, drawing in excess water that stretched the seed coat and pericarp. Loss of water during grain maturation then led to shrinkage when the grains matured. Maltose accumulation is likely to account for the reduced starch synthesis in transgenic grains, through signalling and toxic effects. Using bioinformatics, we identify an isoform of ß-amylase likely to be responsible for maltose accumulation. Removal of this isoform through identification of TILLING mutants or genome editing, combined with co-expression of heterologous glucan synthase and a glucan branching enzyme, may in future enable elevated yields of carbohydrate through simultaneous accumulation of starch and cytosolic glucan.

Restriction of cytosolic sucrose hydrolysis profoundly alters development, metabolism, and gene expression in Arabidopsis roots.

Plant roots depend on sucrose imported from leaves as the substrate for metabolism and growth. Sucrose and hexoses derived from it are also signalling molecules that modulate growth and development, but the importance for signalling of endogenous changes in sugar levels is poorly understood. We report that reduced activity of cytosolic invertase, which converts sucrose to hexoses, leads to pronounced metabolic, growth, and developmental defects in roots of Arabidopsis (Arabidopsis thaliana) seedlings. In addition to altered sugar and downstream metabolite levels, roots of cinv1 cinv2 mutants have reduced elongation rates, cell and meristem size, abnormal meristematic cell division patterns, and altered expression of thousands of genes of diverse functions. Provision of exogenous glucose to mutant roots repairs relatively few of the defects. The extensive transcriptional differences between mutant and wild-type roots have hallmarks of both high sucrose and low hexose signalling. We conclude that the mutant phenotype reflects both low carbon availability for metabolism and growth and complex sugar signals derived from elevated sucrose and depressed hexose levels in the cytosol of mutant roots. Such reciprocal changes in endogenous sucrose and hexose levels potentially provide rich information about sugar status that translates into flexible adjustments of growth and development.

Starch: A Flexible, Adaptable Carbon Store Coupled to Plant Growth.

Research in the past decade has uncovered new and surprising information about the pathways of starch synthesis and degradation. This includes the discovery of previously unsuspected protein families required both for processes and for the long-sought mechanism of initiation of starch granules. There is also growing recognition of the central role of leaf starch turnover in making carbon available for growth across the day-night cycle. Sophisticated systems-level control mechanisms involving the circadian clock set rates of nighttime starch mobilization that maintain a steady supply of carbon until dawn and modulate partitioning of photosynthate into starch in the light, optimizing the fraction of assimilated carbon that can be used for growth. These discoveries also uncover complexities: Results from experiments with Arabidopsis leaves in conventional controlled environments are not necessarily applicable to other organs or species or to growth in natural, fluctuating environments.

Monitoring canid scent marking in space and time using a biologging and machine learning approach.

For canid species, scent marking plays a critical role in territoriality, social dynamics, and reproduction. However, due in part to human dependence on vision as our primary sensory modality, research on olfactory communication is hampered by a lack of tractable methods. In this study, we leverage a powerful biologging approach, using accelerometers in concert with GPS loggers to monitor and describe scent-marking events in time and space. We performed a validation experiment with domestic dogs, monitoring them by video concurrently with the novel biologging approach. We attached an accelerometer to the pelvis of 31 dogs (19 males and 12 females), detecting raised-leg and squat posture urinations by monitoring the change in device orientation. We then deployed this technique to describe the scent marking activity of 3 guardian dogs as they defend livestock from coyote depredation in California, providing an example use-case for the technique. During validation, the algorithm correctly classified 92% of accelerometer readings. High performance was partly due to the conspicuous signatures of archetypal raised-leg postures in the accelerometer data. Accuracy did not vary with the weight, age, and sex of the dogs, resulting in a method that is broadly applicable across canid species' morphologies. We also used models trained on each individual to detect scent marking of others to emulate the use of captive surrogates for model training. We observed no relationship between the similarity in body weight between the dog pairs and the overall accuracy of predictions, although models performed best when trained and tested on the same individual. We discuss how existing methods in the field of movement ecology can be extended to use this exciting new data type. This paper represents an important first step in opening new avenues of research by leveraging the power of modern-technologies and machine-learning to this field.

A comparison of ground-based methods for obtaining large-scale, high-resolution data on the spring leaf phenology of temperate tree species.

Phenological variation in spring leafing between and within species can determine plant responses to warmer winter and spring temperatures in the short term. Methods are needed for monitoring canopy development that can be replicated on a large-scale, while retaining fine-scale resolution at the level of individual trees. Citizen science has the potential to provide this, but a range of approaches exist in terms of the phenophase recorded (e.g. budburst or leaf expansion), how the phenophase is characterised (first events or intensity monitoring) and the portion of tree crown assessed and observation frequency. A comparison of spring budburst and leaf expansion of four tree species (Fraxinus excelsior, Fagus sylvatica, Quercus robur and Acer pseudoplatanus) was monitored in one woodland using (1) counts of expanded leaves on three crown sections, (2) percentage estimates of expanded leaves across the whole crown and (3) a greenness index from photography. Logistic growth models were applied to make comparisons. First-event dates were found to be misleading due to high variation in leaf development rates within and between species. Percentage estimates and counts produced similar estimates of leaf expansion timing and rate. The greenness index produced similar estimates of timing, but not rate, and was compromised by practicalities of photographing individual crowns in closed-canopy woodland. Citizen scientists could collect data across the period of spring leafing, with visual counts and/or estimates made every 3-4 days, subject to tests of reliability in pilot citizen science studies.

Natural Polymorphisms in Arabidopsis Result in Wide Variation or Loss of the Amylose Component of Starch.

Starch granules contain two Glc polymers, amylopectin and amylose. Amylose makes up approximately 10% to 30% (w/w) of all natural starches thus far examined, but mutants of crop and model plants that produce amylose-free starch are generally indistinguishable from their wild-type counterparts with respect to growth, starch content, and granule morphology. Since the function and adaptive significance of amylose are unknown, we asked whether there is natural genetic variation in amylose synthesis within a wild, uncultivated species. We examined polymorphisms among the 1,135 sequenced accessions of Arabidopsis (Arabidopsis thaliana) in GRANULE-BOUND STARCH SYNTHASE (GBSS), encoding the enzyme responsible for amylose synthesis. We identified 18 accessions that are predicted to have polymorphisms in GBSS that affect protein function, and five of these accessions produced starch with no or extremely low amylose (< 0.5% [w/w]). Eight further accessions had amylose contents that were significantly lower or higher than that of Col-0 (9% [w/w]), ranging from 5% to 12% (w/w). We examined the effect of the polymorphisms on GBSS function and uncovered three mechanisms by which GBSS sequence variation led to different amylose contents: (1) altered GBSS abundance, (2) altered GBSS activity, and (3) altered affinity of GBSS for binding PROTEIN TARGETING TO STARCH1-a protein that targets GBSS to starch granules. These findings demonstrate that amylose in leaves is not essential for the viability of some naturally occurring Arabidopsis genotypes, at least over short timescales and under some environmental conditions and open an opportunity to explore the adaptive significance of amylose.

The plastidial pentose phosphate pathway is essential for postglobular embryo development in Arabidopsis.

Large numbers of genes essential for embryogenesis in Arabidopsis encode enzymes of plastidial metabolism. Disruption of many of these genes results in embryo arrest at the globular stage of development. However, the cause of lethality is obscure. We examined the role of the plastidial oxidative pentose phosphate pathway (OPPP) in embryo development. In nonphotosynthetic plastids the OPPP produces reductant and metabolic intermediates for central biosynthetic processes. Embryos with defects in various steps in the oxidative part of the OPPP had cell division defects and arrested at the globular stage, revealing an absolute requirement for the production via these steps of ribulose-5-phosphate. In the nonoxidative part of the OPPP, ribulose-5-phosphate is converted to ribose-5-phosphate (R5P)-required for purine nucleotide and histidine synthesis-and subsequently to erythrose-4-phosphate, which is required for synthesis of aromatic amino acids. We show that embryo development through the globular stage specifically requires synthesis of R5P rather than erythrose-4-phosphate. Either a failure to convert ribulose-5-phosphate to R5P or a block in purine nucleotide biosynthesis beyond R5P perturbs normal patterning of the embryo, disrupts endosperm development, and causes early developmental arrest. We suggest that seed abortion in mutants unable to synthesize R5P via the oxidative part of the OPPP stems from a lack of substrate for synthesis of purine nucleotides, and hence nucleic acids. Our results show that the plastidial OPPP is essential for normal developmental progression as well as for growth in the embryo.

Cas9-mediated mutagenesis of potato starch-branching enzymes generates a range of tuber starch phenotypes.

We investigated whether Cas9-mediated mutagenesis of starch-branching enzymes (SBEs) in tetraploid potatoes could generate tuber starches with a range of distinct properties. Constructs containing the Cas9 gene and sgRNAs targeting SBE1, SBE2 or both genes were introduced by Agrobacterium-mediated transformation or by PEG-mediated delivery into protoplasts. Outcomes included lines with mutations in all or only some of the homoeoalleles of SBE genes and lines in which homoeoalleles carried several different mutations. DNA delivery into protoplasts resulted in mutants with no detectable Cas9 gene, suggesting the absence of foreign DNA. Selected mutants with starch granule abnormalities had reductions in tuber SBE1 and/or SBE2 protein that were broadly in line with expectations from genotype analysis. Strong reduction in both SBE isoforms created an extreme starch phenotype, as reported previously for low-SBE potato tubers. HPLC-SEC and 1 H NMR revealed a decrease in short amylopectin chains, an increase in long chains and a large reduction in branching frequency relative to wild-type starch. Mutants with strong reductions in SBE2 protein alone had near-normal amylopectin chain-length distributions and only small reductions in branching frequency. However, starch granule initiation was enormously increased: cells contained many granules of <4 µm and granules with multiple hila. Thus, large reductions in both SBEs reduce amylopectin branching during granule growth, whereas reduction in SBE2 alone primarily affects numbers of starch granule initiations. Our results demonstrate that Cas9-mediated mutagenesis of SBE genes has the potential to generate new, potentially valuable starch properties without integration of foreign DNA into the genome.

Multiple circadian clock outputs regulate diel turnover of carbon and nitrogen reserves.

Plants accumulate reserves in the daytime to support growth at night. Circadian regulation of diel reserve turnover was investigated by profiling starch, sugars, glucose 6-phosphate, organic acids, and amino acids during a light-dark cycle and after transfer to continuous light in Arabidopsis wild types and in mutants lacking dawn (lhy cca1), morning (prr7 prr9), dusk (toc1, gi), or evening (elf3) clock components. The metabolite time series were integrated with published time series for circadian clock transcripts to identify circadian outputs that regulate central metabolism. (a) Starch accumulation was slower in elf3 and prr7 prr9. It is proposed that ELF3 positively regulates starch accumulation. (b) Reducing sugars were high early in the T-cycle in elf3, revealing that ELF3 negatively regulates sucrose recycling. (c) The pattern of starch mobilization was modified in all five mutants. A model is proposed in which dawn and dusk/evening components interact to pace degradation to anticipated dawn. (d) An endogenous oscillation of glucose 6-phosphate revealed that the clock buffers metabolism against the large influx of carbon from photosynthesis. (e) Low levels of organic and amino acids in lhy cca1 and high levels in prr7 prr9 provide evidence that the dawn components positively regulate the accumulation of amino acid reserves.

Starch granule initiation and morphogenesis-progress in Arabidopsis and cereals.

Starch, the major storage carbohydrate in plants, is synthesized in plastids as semi-crystalline, insoluble granules. Many organs and cell types accumulate starch at some point during their development and maturation. The biosynthesis of the starch polymers, amylopectin and amylose, is relatively well understood and mostly conserved between organs and species. However, we are only beginning to understand the mechanism by which starch granules are initiated, and the factors that control the number of granules per plastid and the size/shape of granules. Here, we review recent progress in understanding starch granule initiation and morphogenesis. In Arabidopsis, granule initiation requires several newly discovered proteins with specific locations within the chloroplast, and also on the availability of maltooligosaccharides which act as primers for initiation. We also describe progress in understanding granule biogenesis in the endosperm of cereal grains-within which there is large interspecies variation in granule initiation patterns and morphology. Investigating whether this diversity results from differences between species in the functions of known proteins, and/or from the presence of novel, unidentified proteins, is a promising area of future research. Expanding our knowledge in these areas will lead to new strategies for improving the quality of cereal crops by modifying starch granule size and shape in vivo.

Final grain weight is not limited by the activity of key starch-synthesising enzymes during grain filling in wheat.

Since starch is by far the major component of the mature wheat grain, it has been assumed that variation in the capacity for starch synthesis during grain filling can influence final grain weight. We investigated this assumption by studying a total of 54 wheat genotypes including elite varieties and landraces that were grown in two successive years in fields in the east of England. The weight, water content, sugars, starch, and maximum catalytic activities of two enzymes of starch biosynthesis, ADP-glucose pyrophosphorylase and soluble starch synthase, were measured during grain filling. The relationships between these variables and the weights and starch contents of mature grains were analysed. Final grain weight showed few or no significant correlations with enzyme activities, sugar levels, or starch content during grain filling, or with starch content at maturity. We conclude that neither sugar availability nor enzymatic capacity for starch synthesis during grain filling significantly influenced final grain weight in our field conditions. We suggest that final grain weight may be largely determined by developmental processes prior to grain filling. Starch accumulation then fills the grain to a physical limit set by developmental processes. This conclusion is in accord with those from previous studies in which source or sink strength has been artificially manipulated.

Leaf Starch Turnover Occurs in Long Days and in Falling Light at the End of the Day.

We investigated whether starch degradation occurs at the same time as starch synthesis in Arabidopsis (Arabidopsis thaliana) leaves in the light. Starch accumulated in a linear fashion for about 12 h after dawn, then accumulation slowed and content plateaued. Following decreases in light intensity, the rate of accumulation of starch declined in proportion to the decline in photosynthesis if the decrease occurred <10 h after dawn, but accumulation ceased or loss of starch occurred if the same decrease in light intensity was imposed more than 10 h after dawn. These changes in starch accumulation patterns after prolonged periods in the light occurred at both high and low starch contents and were not related to time-dependent changes in either the rate of photosynthesis or the partitioning of assimilate between starch and Suc, as assessed from metabolite measurements and 14CO2 pulse experiments. Instead, measurements of incorporation of 13C from 13CO2 into starch and of levels of the starch degradation product maltose showed that substantial starch degradation occurred simultaneously with synthesis at time points >14 h after dawn and in response to decreases in light intensity that occurred >10 h after dawn. Starch measurements in circadian clock mutants suggested that the clock influences the timing of onset of degradation. We conclude that the propensity for leaf starch to be degraded increases with time after dawn. The importance of this phenomenon for efficient use of carbon for growth in long days and for prevention of starvation during twilight is discussed.

Rubisco small subunits from the unicellular green alga Chlamydomonas complement Rubisco-deficient mutants of Arabidopsis.

Introducing components of algal carbon concentrating mechanisms (CCMs) into higher plant chloroplasts could increase photosynthetic productivity. A key component is the Rubisco-containing pyrenoid that is needed to minimise CO2 retro-diffusion for CCM operating efficiency. Rubisco in Arabidopsis was re-engineered to incorporate sequence elements that are thought to be essential for recruitment of Rubisco to the pyrenoid, namely the algal Rubisco small subunit (SSU, encoded by rbcS) or only the surface-exposed algal SSU α-helices. Leaves of Arabidopsis rbcs mutants expressing 'pyrenoid-competent' chimeric Arabidopsis SSUs containing the SSU α-helices from Chlamydomonas reinhardtii can form hybrid Rubisco complexes with catalytic properties similar to those of native Rubisco, suggesting that the α-helices are catalytically neutral. The growth and photosynthetic performance of complemented Arabidopsis rbcs mutants producing near wild-type levels of the hybrid Rubisco were similar to those of wild-type controls. Arabidopsis rbcs mutants expressing a Chlamydomonas SSU differed from wild-type plants with respect to Rubisco catalysis, photosynthesis and growth. This confirms a role for the SSU in influencing Rubisco catalytic properties.