Tumorigenicity of green turtle fibropapilloma-derived fibroblast lines in immunodeficient mice

Fibroblast lines derived from normal skin and spontaneous or experimentally induced fibropapillomas of green turtles (Chelonia mydas) were established and propagated in medium composed of a combination of Dulbecco's minimal essential with F12 medium plus 10% fetal bovine serum at 30 degrees C. Fibropapilloma-derived fibroblasts were indistinguishable from normal skin fibroblasts in vitro. Tumor lines did not exhibit loss of contact inhibition, anchorage independence, or reduced serum requirements. Inoculation of primary and early-passage tumor cells into the medial margin of the pinna of C57BL/6J-nu/nu, C.B17-scid/scid, or NOD-scid/scid mice, however, resulted in fibroma formation, whereas inoculation of normal skin fibroblasts did not. Tumor-derived cells inoculated into the flanks of mice did not form tumors. The turtle origin of fibroblasts in tumors from mouse ears was confirmed by immunohistochemical and karyotype analysis. Fibroblast lines that were established from mouse ear fibromas had the normal karyotype (modal 2N = 55) of C. mydas. The cooler anatomic sites (ears) of immunodeficient mice are useful for confirming the tumorigenic (transformed) phenotype of green turtle fibropapillomatosis-derived fibroblasts. This mouse ear tumorigenicity test should facilitate studies of mechanisms of cellular transformation in green turtle fibropapillomatosis and other neoplastic diseases of poikilothermic vertebrates.     

Effects of surface adsorption on catalytic activity of heavy meromyosin studied using a fluorescent ATP analogue

Biochemical studies in solution and with myosin motor fragments adsorbed to surfaces (in vitro motility assays) are invaluable for elucidation of actomyosin function. However, there is limited understanding of how surface adsorption affects motor properties, e.g., catalytic activity. Here we address this issue by comparing the catalytic activity of heavy meromyosin (HMM) in solution and adsorbed to standard motility assay surfaces [derivatized with trimethylchlorosilane (TMCS)]. For these studies we first characterized the interaction of HMM and actomyosin with the fluorescent ATP analogue adenosine 5'-triphosphate Alexa Fluor 647 2'- (or 3'-) O-(N-(2-aminoethyl)urethane) hexa(triethylammonium) salt (Alexa-ATP). The data suggest that Alexa-ATP is hydrolyzed by HMM in solution at a slightly higher rate than ATP but with a generally similar mechanism. Furthermore, Alexa-ATP is effective as a fuel for HMM-propelled actin filament sliding. The catalytic activity of HMM on TMCS surfaces was studied using (1) Alexa-ATP in total internal reflection fluorescence (TIRF) spectroscopy experiments and (2) Alexa-ATP and ATP in HPLC-aided ATPase measurements. The results support the hypothesis of different HMM configurations on the surface. However, a dominant proportion of the myosin heads were catalytically active, and their average steady-state hydrolysis rate was slightly higher (with Alexa-ATP) or markedly higher (with ATP) on the surface than in solution. The results are discussed in relation to the use of TMCS surfaces and Alexa-ATP for in vitro motility assays and single molecule studies. Furthermore, we propose a novel TIRF microscopy method to accurately determine the surface density of catalytically active myosin motors.     

Lichen respiration in relation to active time, temperature, nitrogen and ergosterol concentrations

1. Respiration in eight lichen species was related to thallus hydration status, external temperature and to total nitrogen, chitin and ergosterol concentrations. Chitin is a nitrogenous and major compound of the fungal cell wall, and ergosterol is a sterol of the plasma membrane in fungi and sometimes in algae.
2. Hydration of previously dry thalli resulted in an initially high rate of respiration. Both the amplitude of this resaturation respiration and the time required to reach steady state varied among species. Generally, peak rates were one to three times higher than steady-state rates, which were reached 3–7 h after hydration.
3. Increases in external temperature also resulted in transient bursts in respiration. Again, both the amplitude of the burst and the time required to reach steady state varied among species. Also depending on species, a temperature increase from 5 to 15 °C resulted in two- to fivefold increases in steady-state respiration.
4. Steady-state respiration, at optimal thallus hydration and a given temperature, varied three- to sixfold among the species, when related to thallus dry mass. This difference correlated best ( r ² = 0·89) with their ergosterol concentration, where a doubling in ergosterol resulted in more than a doubling in respiration. Respiration correlated less well to total nitrogen or chitin.
5. The chitin to ergosterol ratio varied more than one order of magnitude between the species, where species with high nitrogen concentrations had the highest ratio. This implies that species with access to ample amounts of nitrogen can make more fungal cell walls in relation to plasma membrane surface area.     

Adapting to Regional Enforcement: Fishing Down the Governance Index

Background

Illegal, Unreported and Unregulated (IUU) fishing is a problem for marine resource managers, leading to depletion of fish stocks and negative impacts on marine ecosystems. These problems are particularly evident in regions with weak governance. Countries responsible for sustainable natural resource management in the Southern Ocean have actively worked to reduce IUU fishing in the region over a period of 15 years, leading to a sequence of three distinct peaks of IUU fishing.

Methodology/Principal Findings

We reviewed existing public records relating to IUU fishing in the Southern Ocean between 1995–2009 and related this information to the governance capacity of flag states responsible for IUU vessels. IUU operators used a number of methods to adapt to enforcement actions, resulting in reduced risks of detection, apprehension and sanctioning. They changed fishing locations, vessel names and flag states, and ports for offloading IUU catches. There was a significant decrease in the proportion of IUU vessels flagged to CCAMLR countries, and a significant decrease in the average governance index of flag states. Despite a decreasing trend of IUU fishing, further actions are hampered by the regional scope of CCAMLR and the governance capacity of responsible states.

Conclusions/Significance

This is the first study of long-term change in the modus operandi of IUU fishing operators, illustrating that IUU operators can adapt to enforcement actions and that such dynamics may lead to new problems elsewhere, where countries have a limited capacity. This outsourcing of problems may have similarities to natural resource extraction in other sectors and in other regions. IUU fishing is the result of a number of factors, and effectively addressing this major challenge to sustainable marine resource extraction will likely require a stronger focus on governance. Highly mobile resource extractors with substantial funds are able to adapt to changing regulations by exploiting countries and regions with limited capacity.     

Structural basis of T-cell specificity and activation by the bacterial superantigen TSST-1

Superantigens (SAGs) bind simultaneously to major histocompatibility complex (MHC) and T-cell receptor (TCR) molecules, resulting in the massive release of inflammatory cytokines that can lead to toxic shock syndrome (TSS) and death. A major causative agent of TSS is toxic shock syndrome toxin-1 (TSST-1), which is unique relative to other bacterial SAGs owing to its structural divergence and its stringent TCR specificity. Here, we report the crystal structure of TSST-1 in complex with an affinity-matured variant of its wild-type TCR ligand, human T-cell receptor beta chain variable domain 2.1. From this structure and a model of the wild-type complex, we show that TSST-1 engages TCR ligands in a markedly different way than do other SAGs. We provide a structural basis for the high TCR specificity of TSST-1 and present a model of the TSST-1-dependent MHC-SAG-TCR T-cell signaling complex that is structurally and energetically unique relative to those formed by other SAGs. Our data also suggest that protein plasticity plays an exceptionally significant role in this affinity maturation process that results in more than a 3000-fold increase in affinity.     

苹果园光能截获率的数学模型

应用气象学原理推导了不同纬度、不同栽植行向、不同树形的苹果园光能截获率的数学模型,给出在保证树冠基部外围日照时间大于25％总日照时间的条件下,生长季中充分截获光能的最佳树形、行距、冠高、冠径的优化组合方案。实例分析表明,计算结果与专家经验基本一致。本研究为果园合理栽植,充分利用光能,合理整形修剪提供理论依据与参考方案。