Phosphate (Pi) deficiency is a global problem for food production. Plants have evolved complex mechanisms to adapt to low Pi. We focused on the initial aspects of adaptation to low Pi - perception and immediate-early responses to changes in external Pi. To examine whether a labile repressor controls expression of the high affinity Pi transporter, Pht1;1, we performed electrophoretic mobility shift assays (EMSA) but observed only weak protein-DNA binding activity using extracts from Arabidopsis suspension cultures or seedlings. The regulatory role of different regions in Pht1;1 promoter was dissected by promoter deletion analysis, using uidA as a reporter. We identified two domains important for regulation: sequences between -1898 bp and - 932 bp are important for induction of Pht1;1 in low Pi; the intron in the 5’UTR impacts Pht1;1 expression in the young part of both primary and lateral root apices. A complementary approach to identify repressors of Pi starvation responses was pursued: We identified ZAT18, a putative transcription factor, as a candidate repressor. ZAT18 contains an EAR motif, a repressor domain in plants; the expression of ZAT18 responds to Pi starvation. Using transgenic lines with promoter::ZAT18-VENUS constructs, we studied its expression, localization and abundance in different levels of Pi availability: ZAT18 is mainly expressed in the nucleus of Arabidopsis root hair cells. Its accumulation was induced by 4 day Pi starvation. We also performed a microarray analysis to examine global gene expression levels during Pi starvation and rapid recovery. Our data indicated that 258 genes were induced and 188 genes were suppressed during Pi starvation. For most of these genes, responses were reversed after 4 hour Pi recovery. Further study of these genes will help to define targets of the early Pi starvation-signalling pathway.

Sob o efeito do jejum os animais podem se comportar de várias maneiras e mudar a estratégia de forrageio, se arriscar mais ao se alimentar e responder mais rapidamente à presença da presa. Animais ectotérmicos e de metabolismo baixo são bastante afetados pelo jejum, construindo menos abrigos, se alimentando mais e se tornando mais ativos. Escorpiões são predadores predominantemente senta-e-espera mas que podem também deslocar-se em busca de suas presas, especialmente artrópodes. No forrageio, eles dependem de vibrações do substrato e de correntes de ar para localizar e capturar a presa. Esses estímulos são detectados por estruturas como os órgãos em fenda (fina região da cutícula onde ocorrem fendas que detectam vibrações do substrato) e os tricobótrios, que são longas cerdas sensíveis ao deslocamento de ar. Escorpiões são equipados com tricobótrios, mas a função comportamental destas cerdas nesse táxon nunca foi testada experimentalmente. No primeiro capítulo, testamos as hipóteses de que escorpiões em jejum se locomoveriam mais, detectariam mais rapidamente a presa e que a taxa de captura seria maior. Para medir a locomoção, nós dividimos a arena em quadrantes de 11 x 11cm e quantificamos o número de quadrantes utilizados, o número de mudanças de quadrantes e se o animal estava andando ou parado em cada observação. Para quantificar mudanças no limiar de detecção, nós medimos a distância em centímetros e a latência em segundos para detectar a presa. Para medir se houve diferenças na taxa de captura, nós contamos em ambos os grupos o número total de sujeitos experimentais que capturaram o grilo. Nós não encontramos diferenças entre os tratamentos em nenhuma das variáveis medidas. No segundo capítulo, testamos a hipótese de que os tricobótrios são importantes para captura de presas para escorpiões. Previmos que os sujeitos experimentais sem tricobótrios experimentalmente removidos teriam menor sucesso de captura do que os grupos controles (cerda controle removida/controle do procedimento experimental/controle sem manuseio). Previmos ainda que escorpiões sem tricobótrios teriam um maior número de tentativas de captura, que a latência para detecção e para a primeira tentativa de captura seriam maiores e que se orientariam menos vezes para a presa do que animais controle. Colocamos um sujeito experimental e um grilo sem o par de pernas III em uma arena circular com papel sulfite como substrato e filmamos os sujeitos experimentais. Nós não encontramos diferenças nas variáveis medidas entre os grupos comparados. Concluindo, os resultados do primeiro capítulo indicam que talvez o tempo de jejum não tenha sido longo o suficiente para influenciar mudanças no comportamento de forrageio e/ou as diferenças podem aparecer em outros comportamentos não medidos, como o tempo de ingestão. No segundo capítulo, os resultados indicam que os tricobótrios não são essenciais para captura de presas. Dessa forma, outras presas e substratos que atenuam vibrações do substrato devem ser considerados em estudos futuros / Under starvation, animals might behave in different ways and change their foraging strategy, become risk-prone while feeding and respond more quickly to the presence of prey. Ectothermic and with low metabolism animals are greatly affected by starvation, building less burrows, feeding more and becoming more active. Scorpions are predominantly sit-and-wait predators but they may also move in search for prey, especially arthropods. While foraging, they rely on substrate borne vibrations and air displacement to detect and capture prey. These stimuli are detect by sensory structures such as slit sense organs, (a thin region on the cuticle that bears slit that detect substrate borne vibration) and trichobothria, which are long setae sensitive to air displacement. Scorpions are equipped with trichobothria, however, the specific behavioral function of these sensilla on this taxa has never been experimentally tested. In the first chapter, we hypothesized that starved scorpions would become more roving, that the threshold to detection would decrease and that the capture rate would increase. To measure locomotion, we divided the arena into squares of 11 x 11 cm and quantified the number of squares used, the number of square changes and if the animal was walking or staying still in each observation. To quantify changes in the detection threshold, we measured the distance in centimeters and latency in seconds to detect the crickets. To measure if there was a difference in capture rate, we counted in both groups the total number of experimental subjects that captured the cricket. We found no differences between treatments in any of the measured variables. In the second chapter, we tested the hypothesis that trichobothria on pedipalps are important for scorpions to capture prey. We predicted that experimental subjects without trichobothria experimentally removed would be less successful in capturing prey than the control groups (control seta removed/control of experimental procedure/no handling control). We also predicted that scorpions without trichobothria would have a higher number of capture attempts, that the latency to detect prey and to the first capture attempt would be higher and the number of times that each scorpion oriented its body towards the prey would be lower. We used an experimental subject and a cricket without the pair of legs III in a circular arena with a printer paper as substrate and then record the experimental subjects. We did not find differences in the measured variables between the two groups. In conclusion, the results of the first chapter indicate that perhaps the starvation period has not been long enough to influence changes on foraging behavior and/or the differences may appear in other behaviors, such as ingestion time. In the second chapter, the results indicate that trichobothria are not essential for capture prey. Thus, other prey and substrates that attenuate substrate borne vibrations should be considered in future studies

Microalgae hold a great potential to serve as feedstocks for biodiesel production. Tropical and subtropical zones have more solar energy than temperate areas for microalgal culturing. However, outdoor high irradiance results in high temperature in the culturing medium, which is damaging if not lethal to most known microalgal species. New microalgae with heat tolerance, high growth rates and high lipid contents are desirable to establish this industry. Four new green microalgae were isolated in southern Taiwan, located in the subtropical zone. All four species are members of the genus Desmodesmus in the family Senedesmaceae. Two of the four species survived at 45¢Jfor 24 hours, with 5 to 13% mortality rates caused by the heat. Lipid contents of two species reached over 50% in dry biomass under nitrogen starvation. Oil accumulation in tion. Oil accumulation in tion. Oil accumulation in tion. Oil accumulation in the four species positively correlates with their photosystem efficiencies during stress treatments (R2=0.90).

Critical to a cell's survival is its ability to deal with stress by making an appropriate response. This response often takes place in the cytoplasm, which is everything contained within the cell's plasma membrane that is not the nucleus. The cytoplasm is a dynamic environment and its ability to reorganize is essential to the cell's function. This dissertation presents a novel, previously undiscovered state of cytoplasm organization for the model system Schizosaccharomyces pombe, also known as fission yeast. Typically the fission yeast cytoplasm is a fluid-like environment in which endogenous lipid granules subject to thermal fluctuations, move freely as they explore their local surroundings through diffusion. When the cell is in a nutrient depleted environment it is exposed to the stress of advanced glucose starvation. As a result, we find that the cytoplasm undergoes drastic reorganization reminiscent of a phase transition; it is now a solid-like environment in which there is no visible motion. Lipid granules throughout the cell appear to be completely immobilized and are unable to move through the cytoplasm, despite the application of force through optical tweezers. We term this cytoplasmic state the cell frozen state. The cell frozen state is a physiological state, one that the cell can recover from with the addition of fresh nutrients. It is characterized by an anomalous diffusion exponent of [alpha] = 0.23 ± 0.01, which is a significant reduction from the anomalous diffusion exponent [alpha] = 0.66 ± 0.01 found for exponentially growing cells in which there is visible motion. To account for the cell wide immobilization of lipid granules, we hypothesize the formation of a polymer network all through the cytoplasm, and identify septins 1-3 as the most likely filament formers. In addition, we find there is an increase in the number of vacuoles in the cytoplasm during starvation, and propose a vacuole-septin model to describe the cytoplasm reorganization for the cell frozen state. / text

FGF21 is a stress-inducible hormone predominantly secreted from the liver. FGF21 acts as a downstream target gene of hepatic transcription factor PPARα that plays an obligatory role in mediating metabolic adaptation responses to prolonged fasting. However, the physiological roles of FGF21 in regulating glucose homeostasis during adaptive starvation responses and its underlying mechanisms remain unknown. Furthermore, FGF21 is induced by both acute and chronic exercise training in both rodents and human. The physiological significances and the underlying mechanisms for exercise-induced FGF21 production have not been explored so far. Therefore, this study aims to investigate: 1) the mechanisms whereby FGF21 mediates the role of PPARα activation in modulating hepatic gluconeogenesis in response to prolonged fasting; 2) the physiological roles and mechanisms whereby FGF21 regulates exercise capacity and insulin sensitivity via its actions in the muscle during exercise; 3) the roles of FGF receptor-1 (FGFR1) and co-receptor βKlotho in mediating the metabolic effects of FGF21 during fasting and exercise. Our results show thatFGF21 is induced by fasting in the liver and enters into the brain. Both FGF21knockout (KO) mice and PPARα KO mice exhibit severe hypoglycemia and defective hepatic gluconeogenesis during prolonged fasting, and these changes are accompanied by impaired activation of the hypothalamic-pituitary-adrenal (HPA)axis and blunted release of corticosterone from adrenalgland. Moreover, intracerebroventricular injection of recombinant FGF21 reverses fasting hypoglycemia and impairment in hepatic gluconeogenesis by restoring corticosterone production in both FGF21 KO and PPARα KO mice. These effects are abrogated by blockage of hypothalamic FGFR1or by pharmacological inhibition of ERK1/2in the hypothalamus. In addition,FGF21 acts directly on the hypothalamic neurons to activate FGFR1/βKlotho-ERK1/2-CREBsignaling pathway, thereby leading to the transcriptional activation of corticotropin-releasing hormone (CRH) and subsequent activation of the HPA axis. FGF21 is also induced in the liver by exercise and FGF21 KO mice have decreased exercise capacity associated with lower mitochondrial content and thus lower insulin sensitivity compared to wildtype (WT)mice after exercise training.AMPK-PGC-1α signaling pathway is impaired in the muscle of FGF21 KO mice during exercise.FGF21treatmentincreasesmitochondrial content in myotubes, while inhibition of AMPK and knockdown of FGFR1, βKlotho and PGC-1αblocksthis effect. Moreover, FGF21 regulates genes and proteins involved in different steps of the autophagy process and these alterations are mediated by FGFR1/βKlotho-PGC-1α signaling cascade. Furthermore, replenishment of FGF21 reverses the reduced mitochondrial content and insulin sensitivity by restoring PGC-1α expression and subsequent increase in mitochondrial biogenesis and suppression of autophagy in the muscle of FGF21 KO mice. In conclusion, our results identify FGF21 as a critical hormonal regulator of glucose homeostasis during prolonged fasting, by coupling hepatic PPARα activation to corticosterone release via stimulation of the HPA axis in the brain. Moreover, FGF21 acts in the muscle through two distinctive mechanism to maintain mitochondrial homeostasis during exercise training, involving both biogenesis of new mitochondria and decreased clearance of stressed mitochondria and thus to mediate the beneficial effects of exercise on lipid and glucose metabolism. / published_or_final_version / Medicine / Doctoral / Doctor of Philosophy

S. cerevisiae have developed the ability to forage for nutrients when presented with conditions of starvation. This dimorphic adaptation is particularly noticeable when yeast are subject to nitrogen depravation and has been termed filamentous growth, as cells form filament-like projections away from the center of the colony. The regulation of this response is under the control of the well-characterized MAPK and cAMP pathways. Previous work showed that Cdk8p phosphorylated a key transcriptional activator of the filamentous response, Ste12p, and subsequently targeted the factor for degradation under conditions of limiting nitrogen. Data presented in this thesis suggests that Cdk8p is regulated by another kinase, Tpk2p. In vitro kinase assays demonstrate that Tpk2p directly phosphorylates Cdk8p on residue Thr37, leading to the destabilization of Cdk8p after growth for 4 hours in SLAD media. Lack of phosphorylation on Thr37 yields a hypo-hypofilamentous phenotype, whereas a phospho-mimic mutant, T37E displays a filamentous hyper-filamentous phenotype.

During nutrient-replete growth of microalgae, new photosynthate is allocated toward three different biochemical pools: light harvesting compounds, the biosynthetic apparatus and energy storage. The mechanisms governing allocation of photosynthate between the energy storage compounds carbohydrate and lipid are not well understood. For biofuel production, it is desirable to identify conditions and algal strains that allocate maximum amounts of photosynthate to lipid. This thesis assessed the allocation of photosynthate toward the energy storage pool, and to lipid vs. carbohydrate, at two light levels and during ongoing nitrogen-starvation, for two diatoms and two chlorophytes. Nitrogen-starvation resulted in an increase in the photosynthate allocated toward energy storage, however the magnitude of change was determined by a combination of species and light level. Of the four species studied, the diatom Chaetoceros muelleri, grown in high light, accumulated lipid during N-starvation at a relatively high rate, making it a good candidate for biofuel production.

Porcine Periweaning Failure-to-Thrive syndrome (PFTS) is a clinical syndrome of newly weaned pigs with unknown etiology and characterized by anorexia, lethargy and progressive debilitation. The hypothesis of this thesis is that PFTS is an infectious disease. Investigation in an index farm affected by PFTS from Saskatchewan Canada ruled out most common swine pathogens as the etiology and identified several lesions that were consistent across many cases. A larger study including multiple farms in North America was then undertaken. A total of 8 farms were investigated, within which 5 met the clinical definition of PFTS. Gross and histological examinations were performed on 8 case and 4 control pigs on each farm. Detection of relevant porcine pathogens, complete blood count, serum chemistry, and serum cytokine analysis were performed on each pig. Thymic atrophy, superficial gastritis and small intestinal villous atrophy were significantly more prevalent in case pigs compared to control pigs. All case pigs had at least two of these three lesions. All case and control pigs were negative for porcine reproductive and respiratory syndrome virus, swine influenza virus and were free of porcine circovirus associated diseases. Although several pathogens, such as porcine cytomegalovirus, haemagglutinating encephalomyelitis virus, porcine enteric calicivirus, group A rotavirus, enteroviruses and Cystoisospora suis were detected in some of the case and control pigs, none were associated with clinical status. Clinical pathology findings of case pigs was consistent with anorexia and dehydration, such as increases in haematocrit, blood urea, serum bilirubin, albumin, beta-hydroxybutyrate and decreases in blood glucose, calcium and phosphorous. Case pigs had similar levels to IL1-β than control pigs, which suggested that PFTS was not a result of excessive cytokines. In subsequent experiments, a snatched-farrowed porcine-colostrum-deprived (SF-pCD) pig model was developed and tissue homogenates were used to inoculate SF-pCD pigs in an attempt to reproduce the clinical signs of PFTS. The SF-pCD pigs were immunologically characterized and shown to be suitable for inoculation studies. However, inoculation of tissue homogenate from PFTS pigs failed to reproduce the clinical signs of PFTS in SF-pCD pigs. All together, PFTS is a clinical syndrome with consistent pathological and serum analytical changes among affected pigs. Despite the efforts of this research to establish an infectious etiology, there is a lack of evidence that PFTS is an infectious disease.

No description available.

The first year of life is one of the main survival bottlenecks for many fish species. Individual traits affecting survival can be morphological, physiological or behavioural. Body size, growth rate and activity have all been found to affect fitness in different organisms. However, the effects of these traits on fitness in natural conditions and for underyearlings are poorly investigated. In this study we attempted to induce compensatory growth in laboratory conditions in natural populations of brown trout fry (Salmo trutta). It was performed by exposing the fry to a period of restricted resources followed by a period of refeeding. Two behavioural trials were conducted on each individual where activity level was scored. All fish were subsequently released in their native stream and recaptured after a month to check for survival. We found that high individual activity level in an open field context increased the probability of survival under natural conditions. The importance of body size for survival decreased over time, and thus, with fish size. Full compensation was detected in body condition, while only partly compensation in weight and no compensation in length were detected during the experimental periods. Our results suggest that a brown trout fry’s individual activity level is repeatable and can be an important trait for selection in nature. The instable interactions between activity and life-history traits indicate environmental effects on these interactions. Furthermore, if body size is not the only trait affecting survival, compensation in body structures may not be a fast response to increase fitness after a period of growth depression.