Environmental sensing by African trypanosomes

African trypanosomes, which divide their life cycle between mammals and tsetse flies, are confronted with environments that differ widely in temperature, nutrient availability and host responses to infection. In particular, since trypanosomes cannot predict when they will be transmitted between hosts, it is vital for them to be able to sense and adapt to their milieu. Thanks to technical advances, significant progress has been made in understanding how the parasites perceive external stimuli and react to them. There is also a growing awareness that trypanosomes use a variety of mechanisms to exchange information with each other, thereby enhancing their chances of survival.

Targeting AKT/mTOR Signaling Pathways During Murine Skin Tumor Promotion and the Impact of Dietary Energy Balance Manipulation

The prevalence of obesity has continued to rise over the last several decades in the United States lending to overall increases in risk for chronic diseases including many types of cancer. In contrast, reduction in energy consumption via calorie restriction (CR) has been shown to be a potent inhibitor of carcinogenesis across a broad range of species and tumor types. Previous data has demonstrated differential signaling through Akt and mTOR via the IGF-1R and other growth factor receptors across the diet-induced obesity (DIO)/CR spectrum. Furthermore, mTORC1 is known to be regulated directly via nutrient availability, supporting its role in the link between epithelial carcinogenesis and diet-induced obesity. In an effort to better understand the importance of mTORC1 in the context of both positive and negative energy balance during epithelial carcinogenesis, we have employed the use of specific pharmacological inhibitors, rapamycin (mTORC1 inhibitor) and metformin (AMPK activator) to target mTORC1 or various components of this pathway during skin tumor promotion. Two-stage skin carcinogenesis studies demonstrated that mTORC1 inhibition via rapamycin, metformin or combination treatments greatly inhibited skin tumor development in normal, overweight and obese mice. Furthermore, mechanisms by which these chemopreventive agents may be exerting their anti-tumor effects were explored. In addition, the effect of these compounds on the epidermal proliferative response was analyzed and drastic decreases in epidermal hyperproliferation and hyperplasia were found. Rapamycin also inhibited dermal inflammatory cell infiltration in a dose-dependent manner. Both compounds also blocked or attenuated TPA-induced signaling through epidermal mTORC1 as well as several downstream targets. In addition, inhibition of this pathway by metformin appeared to be, at least in part, dependent on AMPK activation in the skin. Overall, the data indicate that pharmacological strategies targeting this pathway offset the tumor-enhancing effects of DIO and may serve as possible CR mimetics. They suggest that mTORC1 contributes significantly to the process of skin tumor promotion, specifically during dietary energy balance effects. Exploiting the mechanistic information underlying dietary energy balance responsive pathways will help translate decades of research into effective strategies for prevention of epithelial carcinogenesis.

BIOCHEMICAL CHARACTERIZATION OF BINDING PARTNERS OF TWO HSP70 CO-CHAPERONES IN SACCHAROMYCES CEREVISIAE

Cells are exposed to a variety of environmental and physiological changes including temperature, pH and nutrient availability. These changes cause stress to cells, which results in protein misfolding and altered cellular protein homeostasis. How proteins fold into their three-dimensional functional structure is a fundamental biological process with important relevance to human health. Misfolded and aggregated proteins are linked to multiple neurodegenerative diseases, cardiovascular disease and cystic fibrosis. To combat proteotoxic stress, cells deploy an array of molecular chaperones that assist in the repair or removal of misfolded proteins. Hsp70, an evolutionarily conserved molecular chaperone, promotes protein folding and helps maintain them in a functional state. Requisite co-chaperones, including nucleotide exchange factors (NEFs) strictly regulate and serve to recruit Hsp70 to distinct cellular processes or locations. In yeast and human cells, three structurally non-related cytosolic NEFs are present: Sse1 (Hsp110), Fes1 (HspBP1) and Snl1 (Bag-1). Snl1 is unique among the cytosolic NEFs as it is localized at the ER membrane with its Hsp70 binding (BAG) domain exposed to the cytosol. I discovered that Snl1 distinctly interacts with assembled ribosomes and several lines of evidence indicate that this interaction is both independent of and concurrent with binding to Hsp70 and is not dependent on membrane localization. The ribosome-binding site is identified as a short lysine-rich motif within the amino terminus of the Snl1 BAG domain distinct from the Hsp70 interaction region. In addition, I demonstrate ribosome association with the Snl1 homolog in the pathogenic fungus, Candida albicans and localize this putative NEF to a perinuclear/ER membrane, suggesting functional conservation in fungal BAG domain-containing proteins. As a first step in determining specific domain architecture in fungal BAG proteins, I present the preliminary steps of protein purification and analysis of the minimal Hsp70 binding region in in both S.cerevisiae and C. albicans Snl1. Contrary to previous in vitro evidence which showed the Fes1 NEF to interact with both cytosolic Hsp70s, Ssa and Ssb, Fes1 is shown to interact specifically with Ssa when expressed under normal cellular conditions in S. cerevisiae. This is the first reported evidence of Hsp70 binding selectivity for a cytosolic NEF, and suggests a possible mechanism to achieve specificity in Hsp70-dependent functions. Taken together, the work presented in this dissertation highlights the striking divergence among Hsp70 co-chaperones in selecting binding partners, which may correlate with their specific roles in the cell.

Siliceous microfossils in middle Eocene sediments of ODP Holes 120-748B and 120-749B

The Middle Eocene Climatic Optimum (MECO) is a major transient warming event that occurred at ~ 40 Ma and reversed a long-term cooling trend through the early and middle Eocene. We report the results of a high-resolution, quantitative study of siliceous microfossils at Ocean Drilling Program Sites 748 and 749 (Southern Kerguelen Plateau, Southern Ocean, ~ 58°S) across a ~ 1.4 myr interval spanning the MECO event. At both sites, a significant increase in biosiliceous sedimentation is associated with the MECO event. Rich siliceous planktonic microfossil assemblages in this interval are unusual in that they are dominated by ebridians, with radiolarians as a secondary major component. Silicoflagellates and diatoms comprise only a minor fraction of the assemblage, in contrast to siliceous microfossil assemblages that characterize modern Southern Ocean sediments. Based on our new siliceous microfossil records, we interpret two ~ 300 kyr periods of elevated nutrient availability in Southern Ocean surface waters which span the peak warming interval of the MECO and the post-MECO cooling interval. A diverse assemblage of large silicoflagellates belonging to the Dictyocha grandis plexus is linked to the rapid rise in sea-surface temperatures immediately prior to peak warmth, and a pronounced turnover is observed in both ebridian and silicoflagellate assemblages at the onset of peak warming. The interval of peak warmth is also characterized by high abundance of cosmopolitan ebridians (e.g., Ammodochium spp.) and silicoflagellates (e.g., Naviculopsis spp.), and increased abundance of tropical and subtropical diatom genera (e.g., Asterolampra and Azpeitia). These observations confirm the relative pattern of temperature change interpreted from geochemical proxy data at multiple Southern Ocean sites. Furthermore, rapid assemblage changes in both autotrophic and heterotrophic siliceous microfossil groups indicate a reorganization of Southern Ocean plankton communities in response to greenhouse warming during the MECO event.

Fertilización nitrogenada en plantines de pimiento (Capsicum annuum L.) cultivados en sustratos con diferentes proporciones de materiales compostados : efecto sobre los parámetros de calidad del plantín

La etapa inicial en el crecimiento de los plantines constituye el momento más crítico en su producción. Los materiales compostados pueden resultar beneficiosos por el aporte de nutrientes y la mejora en la condición física del medio de crecimiento. Para evaluar el efecto de sustratos preparados con y sin materiales compostados [Testigo (60% turba de Sphagnum+40% perlita); Mezcla I (45% turba de Sphagnum+30% perlita+25% material compostado); Mezcla II (30% turba de Sphagnum+20% perlita+50% material compostado) y un sustrato Comercial (turba de Sphagnum+40% compost+perlita+vermiculita], sobre la nutrición inicial de plantines de pimiento (Capsicum annuum L.), se realizó un ensayo fertilizando con 0, 150 y 300 mg N L-1. Se determinaron altura, diámetro del tallo y pesos frescos y secos de hoja, tallo y raíz. Se calculó relación vástago/raíz y hoja/tallo en fresco y seco, y porcentual de materia seca. El diseño experimental fue completamente aleatorizado con cuatro repeticiones. Los materiales compostados mejoraron la calidad de los plantines, los cuales no fueron afectados cuando se aplicó N a los sustratos con compost, y sólo se observaron leves mejoras en el crecimiento de los plantines al fertilizar los sustratos carentes de compost, debido a su escasa retención hídrica y elevada lixiviación de N.

Sea surface reconstruction for sediments of the California continental margin

Organic matter in sediment samples from three ODP sites (Ocean Drilling Program Leg 167) that form a south-north transect was investigated to reconstruct the paleoclimatic and oceanographic conditions on the California continental margin during the last 160 kyr. Alkenone-derived paleosea surface temperatures (SST) are 3 to 6°C colder in glacial stages and reveal a clear relationship with global climate changes; the differences are greater in the north. Latitudinal SST comparison exhibits water mixing of the colder California Current with warmer waters from the south, particularly in the southern central California borderland area. Organic matter accumulation on the California continental margin indicates an interplay between climatic and atmospheric glacial-interglacial variations and spatially and temporally changing nutrient availability along the California coastline. Climatic and atmospheric dependent circulations apparently caused variations in the intensity of coastal upwelling along the southern central California margin and this suggests, due to the close connection of the California Current to the local wind patterns, that the California Current was weaker during glacial and stronger during interglacial periods.

Geochemistry on Holocene sediments from the Mediterranean

The enhanced accumulation of organic matter in Eastern Mediterranean sapropels and their unusually low d15N values have been attributed to either enhanced nutrient availability which led to elevated primary production and carbon sequestration or to enhanced organic matter preservation under anoxic conditions. In order to evaluate these two hypothesis we have determined Ba/Al ratios, amino acid composition, N and organic C concentrations and d15N in sinking particles, surface sediments, eight spatially distributed core records of the youngest sapropel S1 (10-6 ka) and older sapropels (S5, S6) from two locations. These data suggest that (i) temporal and spatial variations in d15N of sedimentary N are driven by different degrees of diagenesis at different sites rather than by changes in N-sources or primary productivity and (ii) present day TOC export production would suffice to create a sapropel like S1 under conditions of deep-water anoxia. This implies that both enhanced TOC accumulation and d15N depletion in sapropels were due to the absence of oxygen in deep waters. Thus preservation plays a major role for the accumulation of organic-rich sediments casting doubt on the need of enhanced primary production for sapropel formation.

Physical, chemical and biological characteristics of disturbed and undisturbed Arctic lakes, Northwest Territories

1. Global warming is predicted to cause changes in permafrost cover and stability in the Arctic. Zones of high ion concentration in regions of ice-rich permafrost are a reservoir of chemicals that can potentially be transferred to fresh waters during thawing. Consequently, input of enriched runoff from the thaw and sediment and vegetation from the landscape could alter lakes by affecting their geochemistry and biological production. 2. Three undisturbed lakes and five lakes disturbed by retrogressive permafrost thaw slumps were sampled during late summer of 2006 to assess the potential effects of thermokarst shoreline slumping on water and sediment chemistry, the underwater light regime, and benthic macrophyte biomass and community structure. 3. Undisturbed lakes had sediments rich in organic material and selected micronutrients, while disturbed lakes had sediments richer in calcium, magnesium and strontium, greater transparency of the water column, and a well-developed submerged macrophyte community. 4. It is postulated that enriched runoff chemistry may alter nutrient availability at the sediment-water interface and also the degradation of organic material, thus affecting lake transparency and submerged macrophytes. The results suggest that retrogressive permafrost slumping can significantly affect food webs in arctic tundra lakes through an increase in macrophyte biomass and development of a more complex benthic habitat.

Planktonic assemblages during sapropel S5 deposits in the Urania Basin

A micropaleontological study of planktonic assemblages on the partially laminated sapropel S5 (late Pleistocene, marine isotope stage (MIS) 5e) was performed in two piston cores from Urania Basin area (eastern Mediterranean, west of Crete): UM94PC16 and UM94PC31 recovered during a PALEOFLUX Project Cruise. The abundance of Florisphaera profunda indicates the development of a deep chlorophyll maximum (DCM) before the anoxic condition at bottom were established, whereas patterns of upper photic zone coccoliths suggest extreme oligotrophy in surface water. The short appearance of Globorotalia scitula and the presence of Globigerinoides ruber in the lower part of sapropel testify to a thermal stratification, also recorded by changes in primary producers. During G. scitula occurrence, diatoms, mainly represented by Pseudosolenia calcar-avis, appear and bloom because of their capability in using nutrients from DCM. Scanning electron microscope analyses performed on selected intervals from UM94PC16 show that the sapropel is organized in microlaminae mostly composed by siliceous microfossils. In particular, sapropel S5 could be related to an enhanced nutrient availability in the lower-middle part of the photic zone, stratified conditions, and a higher continental input.

Particle flux studies of sediment traps from the northern and equatorial Indian Ocean

The Asian monsoon system governs seasonality and fundamental environmental characteristics in the study area from which two distinct peculiarities are most notable: upwelling and convective mixing in the Arabian Sea and low surface salinity and stratification in the Bay of Bengal due to high riverine input and monsoonal precipitation. The respective oceanography sets the framework for nutrient availability and productivity. Upwelling ensures high nitrate concentration with temporal/spatial Si limitation; freshwater-induced stratification leads to reduced nitrogen input from the subsurface but Si enrichment in surface waters. Ultimately, both environments support high abundance of diatoms, which play a central role in the export of organic matter. It is speculated that, additional to eddy pumping, nitrogen fixation is a source of N in stratified waters and contributes to the low-d15N signal in sinking particles formed under riverine impact. Organic carbon fluxes are best correlated to opal but not to carbonate, which is explained by low foraminiferal carbonate fluxes within the river-impacted systems. This observation points to the necessity of differentiating between carbonate sources for carbon flux modeling. As evident from a compilation of previously published and new data on labile organic matter composition (amino acids and carbohydrates), organic matter fluxes are mainly driven by direct input from marine production, except the site off Pakistan where sedimentary input of (marine) organic matter is dominant during the NE monsoon. The explanation of apparently different organic carbon export efficiency calls for further investigations of, for example, food web structure and water column processes.

Planktic foraminiferal sieve size specific d13C and d18O values and maximum test diameter from ODP sites 171-1051 and 120-748

Many genera of modern planktic foraminifera are adapted to nutrient-poor (oligotrophic) surface waters by hosting photosynthetic symbionts, but it is unknown how they will respond to future changes in ocean temperature and acidity. Here we show that ca. 40 Ma, some fossil photosymbiont-bearing planktic foraminifera were temporarily 'bleached' of their symbionts coincident with transient global warming during the Middle Eocene Climatic Optimum (MECO). At Ocean Drilling Program (ODP) Sites 748 and 1051 (Southern Ocean and mid-latitude North Atlantic, respectively), the typically positive relationship between the size of photosymbiont-bearing planktic foraminifer tests and their carbon isotope ratios (d13C) was temporarily reduced for ~100 k.y. during the peak of the MECO. At the same time, the typically photosymbiont-bearing planktic foraminifera Acarinina suffered transient reductions in test size and relative abundance, indicating ecological stress. The coincidence of minimum d18O values and reduction in test size-d13C gradients suggests a link between increased sea-surface temperatures and bleaching during the MECO, although changes in pH and nutrient availability may also have played a role. Our findings show that host-photosymbiont interactions are not constant through geological time, with implications for both the evolution of trophic strategies in marine plankton and the reliability of geochemical proxy records generated from symbiont-bearing planktic foraminifera.

Benthic foraminifera abundance data of sediment core GeoB7926-2

We present a high resolution, multiproxy study of the relationship between pelagic and benthic environments of a coastal upwelling system in the subtropical NE Atlantic Ocean. Marine sediments corresponding to late MIS3 to the Holocene in the radiocarbon dated core GeoB7926, retrieved off Mauritania (21°N) were analysed to reconstruct productivity in surface waters and its linkage to deep waters during the last 35 ka BP. High latitude cold events and changes in atmospheric and oceanographic dynamics influenced upwelling intensity over this time period. Subsequently, this caused changes in primary productivity off this low-latitude coastal upwelling locality. The benthic foraminiferal fauna displays four main community shifts corresponding to fundamental climatic events, first of all during late MIS3 (35-28 ka BP), secondly from 28 to 19 ka BP (including Heinrich event 2 and the LGM), thirdly within Heinrich event 1, the Bølling Allerød and the Younger Dryas (18-11.5 ka BP) and finally during the Holocene (11.5-0 ka BP). In particular, strong pelagic-benthic coupling is apparent in MIS 3, as demonstrated by increased primary productivity, indicated by moderate DAR and the dominance of benthic foraminiferal species which prefer fresh phytodetritus. A decline in upwelling intensity and nutrient availability follows, which resulted in a proportionately larger amount of older, degraded matter, provoking a shift in the benthic foraminifera fauna composition. This rapid response of the benthic environment continues with a progressive increase in upwelling intensity due to sea level and oceanographic changes and according high surface production during the LGM. During Heinrich event 1 and the Younger Dryas, extreme levels of primary production actually hindered benthic environment through the development of low oxygen conditions. After this period, a final change in benthic foraminiferal community composition occurs which indicates a return to more oxygenated conditions during the Holocene.

Temperature proxies of sediment cores from the Sea of Marmara

Reconstructing ocean temperature values is a major target in paleoceanography and climate research. However, most temperature proxies are organism-based and thus suffer from an "ecological bias". Multiproxy approaches can potentially overcome this bias but typically require more investment in time and resources, while being susceptible to errors induced by sample preparation steps necessary before analysis. Three lipid-based temperature proxies are widely used: UK'37 (based on long chain alkenones from phytoplanktonic haptophytes), TEX86 [based on glycerol dialkyl glycerol tetraethers (GDGTs) from pelagic archaea] and LDI (based on long chain diols from phytoplanktonic eustigmatophytes). So far, separate analytical methods, including gas chromatography (GC) and liquid chromatography (LC), have been used to determine these proxies. Here we present a sensitive method for determining all three in a single normal phase high performance LC-atmospheric pressure chemical ionization mass spectrometry (NP-HPLC-APCI-MS) analysis. Each of the long chain alkenones and long chain diols was separated and unambiguously identified from the accurate masses and characteristic fragmentation during multiple stage MS analysis (MS2). Comparison of conventional GC and HPLC-MS methods showed similar results for UK'37 and LDI, respectively, using diverse environmental samples and an Emiliania huxleyi culture. Including the three sea surface temperature (SST) proxies; the NP-HPLC-APCI-MS method in fact allows simultaneous determination of nine paleoenvironmental proxies. The extent to which the ecology of the source organisms (ecological bias) influences lipid composition and thereby the reconstructed temperature values was demonstrated by applying the new method to a sediment core from the Sea of Marmara, covering the last 21 kyr BP. Reconstructed SST values differed considerably between the proxies for the Last Glacial Maximum (LGM) and the period of Sapropel S1 formation at ca. 10 kyr BP, whereas the trends during the late Holocene were similar. Changes in the composition of alkenone-producing species at the transition from the LGM to the Bølling/Allerød (B/A) were inferred from unreasonably high UK'37-derived SST values (ca. 20 °C) during the LGM. We ascribe discrepancies between the reconstructed temperature records during S1 deposition to habitat change, e.g. a different depth due to changes in nutrient availability.

Calcareous nannofossil assemblages of Ocean Anoxic Event 2 in ODP Sites 207-1258 and 207-1260

The calcareous nannofossils of the Cenomanian/Turonian boundary interval of Sites 1258 and 1260 (Ocean Drilling Program Leg 207) have been studied in order to understand the depositional environment during Oceanic Anoxic Event 2 (OAE2) in the equatorial Atlantic. Nannofossil assemblages show a significant change in relative abundances during the positive d13Corg excursion interval. The strong increase of the high productivity indicator Zeugrhabdotus erectus and the simultaneous decrease of the oligotrophic taxa Watznaueria barnesiae and Watznaueria fossacincta are indicative of enhanced fertility. The decrease of Eprolithus floralis may be attributed to the surface-water temperature increase during OAE2, which is, however, not very significant (~2-3 °C), as suggested by published TEX86 data. It seems more likely that the decrease of E. floralis during OAE2 was evoked by the breakdown of water-column stratification, indicating it as a deep-dwelling species, which prefers stratified waters with a deep nutricline. Prediscosphaera spp. and Retecapsa ficula, which show a significant increase in relative abundances during OAE2, seem to prefer eutrophic environments, while Amphizygus brooksii and Zeugrhabdotus noeliae lower surface-water fertility. Gartnerago segmentatum, Broinsonia spp., Watznaueria biporta, and Seribiscutum gaultensis decrease in abundances during OAE2. It is not clear if they preferred an oligotrophic environment, cooler surface-waters, or if they were inhabitants of the lower photic zone. Published geochemical data suggest that enhanced fertility and higher temperatures during OAE2 may have been caused by submarine volcanic activity through the release of biolimiting micronutrients into the ocean and carbon dioxide into the atmosphere. The breakdown of water-column stratification may have increased further nutrient availability.

Carbon/Nitrogen ratio of plant tissues and biomass of belowground consumers after 11 years of experimental treatment, Toolik lake

Theory and observation indicate that changes in the rate of primary production can alter the balance between the bottom-up influences of plants and resources and the top-down regulation of herbivores and predators on ecosystem structure and function. The Exploitation Ecosystem Hypothesis (EEH) posited that as aboveground net primary productivity (ANPP) increases, the additional biomass should support higher trophic levels. We developed an extension of EEH to include the impacts of increases in ANPP on belowground consumers in a similar manner as aboveground, but indirectly through changes in the allocation of photosynthate to roots. We tested our predictions for plants aboveground and for phytophagous nematodes and their predators belowground in two common arctic tundra plant communities subjected to 11 years of increased soil nutrient availability and/or exclusion of mammalian herbivores. The less productive dry heath (DH) community met the predictions of EEH aboveground, with the greatest ANPP and plant biomass in the fertilized plots protected from herbivory. A palatable grass increased in fertilized plots while dwarf evergreen shrubs and lichens declined. Belowground, phytophagous nematodes also responded as predicted, achieving greater biomass in the higher ANPP plots, whereas predator biomass tended to be lower in those same plots (although not significantly). In the higher productivity moist acidic tussock (MAT) community, aboveground responses were quite different. Herbivores stimulated ANPP and biomass in both ambient and enriched soil nutrient plots; maximum ANPP occurred in fertilized plots exposed to herbivory. Fertilized plots became dominated by dwarf birch (a deciduous shrub) and cloudberry (a perennial forb); under ambient conditions these two species coexist with sedges, evergreen dwarf shrubs, and Sphagnum mosses. Phytophagous nematodes did not respond significantly to changes in ANPP, although predator biomass was greatest in control plots. The contrasting results of these two arctic tundra plant communities suggest that the predictions of EEH may hold for very low ANPP communities, but that other factors, including competition and shifts in vegetation composition toward less palatable species, may confound predicted responses to changes in productivity in higher ANPP communities such as the MAT studied here.