The role of ecosystem transpiration in creating alternate moisture regimes by influencing atmospheric moisture convergence

The terrestrial water cycle links the soil and atmosphere moisture reservoirs through four fluxes: precipitation, evaporation, runoff, and atmospheric moisture convergence (net import of water vapor to balance runoff). Each of these processes is essential for sustaining human and ecosystem well-being. Predicting how the water cycle responds to changes in vegetation cover remains a challenge. Recently, changes in plant transpiration across the Amazon basin were shown to be associated disproportionately with changes in rainfall, suggesting that even small declines in transpiration (e.g., from deforestation) would lead to much larger declines in rainfall. Here, constraining these findings by the law of mass conservation, we show that in a sufficiently wet atmosphere, forest transpiration can control atmospheric moisture convergence such that increased transpiration enhances atmospheric moisture import and results in water yield. Conversely, in a sufficiently dry atmosphere increased transpiration reduces atmospheric moisture convergence and water yield. This previously unrecognized dichotomy can explain the otherwise mixed observations of how water yield responds to re-greening, as we illustrate with examples from China's Loess Plateau. Our analysis indicates that any additional precipitation recycling due to additional vegetation increases precipitation but decreases local water yield and steady-state runoff. Therefore, in the drier regions/periods and early stages of ecological restoration, the role of vegetation can be confined to precipitation recycling, while once a wetter stage is achieved, additional vegetation enhances atmospheric moisture convergence and water yield. Recent analyses indicate that the latter regime dominates the global response of the terrestrial water cycle to re-greening. Evaluating the transition between regimes, and recognizing the potential of vegetation for enhancing moisture convergence, are crucial for characterizing the consequences of deforestation as well as for motivating and guiding ecological restoration.     

Patterns of Integration and Expression of Retroviral, Non-Replicative Vectors in Avian Embryos: Embryo Developmental Stage and Virus Subgroup Envelope Modulate Tissue-Tropism

We previously demonstrated that Avian Leukemia Viruses (ALV) carrying the v-myc gene specifically induce two types of tumors, cardiomyocytic tumors when the virus is injected before embryonic day 3 (E3), skin tumors when the virus is injected at E3 or E5.

Aiming to elucidate the mechanisms which determine this time-dependent change in target, we infected chick and quail embryos at E3 and E5 with replication-deficient, lacZ gene-carrying, ALV-based viruses produced by a packaging cell line. Three constructs driven by 3 different Long Terminal Repeats (LTRs) were tested and yielded similar results. When the constructs were inoculated at E3 and the lacZ gene product revealed 5 days later, around 70% of the embryos carried lacZ+ clones in the heart, around 50% had positive clones in the skin anywhere on the body, while a few embryos displayed clones in internal organs (liver, stomach, lungs). Immunocytological identification of the heart cell type(s) expressing the virus revealed that the only cells infected were cardiomyocytes. When the constructs were inoculated at E5, no lacZ+ clones appeared in the heart but all were located in the cephalic skin. In order to examine the relationship between viral integration and expression, DNA of different organs or tissues from lacZ stained embryos was analyzed by PCR. A tight correlation between integration and expression in the heart and in the skin was revealed in most cases. In contrast, a significant PCR signal was often detected in the liver or the stomach despite weak or absent expression as revealed by lacZ+ clones.

We then investigated the influence of envelope glycoprotein subgroups on the tropism of these constructs. The lacZ vector driven by RAV-2 LTRs was packaged as subgroups A, B or E viral particles. The A subgroup, used in the part of the study described above, infects both chick and quail while the B and E subgroups are specific for chick or quail respectively. These B and E subgroups induced lacZ+ clones in the heart (after E3 injection) while no clones or only a few were detected in the skin either after E3 or E5 injection. The following conclusions can be drawn: 1) cardiomyocytes are at E3 the major target for integration and expression of ALV-derived viruses in vivo; 2) targets change rapidly with embryonic age; and 3) tissue-specific infections depend on the envelope subgroup, thus presumably on the presence of the cognate receptor. This study clearly indicates that E3 inoculation of ALV-based retroviral vectors is a simple and powerful method to transfer gene sequences into cardiomyocytes and epidermal cells.     

Expression of the α-thionin gene from barley in tobacco confers enhanced resistance to bacterial pathogens

Thionins are cysteine-rich, 5 kDa polypeptides which are toxic to plant pathogens in vitro. Expression of the gene encoding α-thionin from barley endosperm, under the 35S promoter from cauliflower mosaic virus, conferred to transgenic tobacco enhanced resistance to the bacterial plant pathogens Pseudomonas syringae pv. tabaci 153 and P. syringae pv. syringae. The barley α-thionin gene, which has two introns, was correctly spliced in tobacco. The α-thionin in transgenic plants had the expected mobility in the gradient, when separated by high-performance liquid chromatography, reacted with monospecific antibodies and showed the expected antibiotic properties in vitro.     

Low-cost method for the preservation of Sclerotium rolfsii Proimi F-6656: Inoculum standardization and its use in scleroglucan production

Summary The Castellani's Method for the preservation of Sclerotium rolfsii in sterile distilled water was tested. Culturing on Potato Dextrose Yeast extract (PDY) slants, the current system used, was also evaluated. Preservation of sclerotia according to the Castellani's method allowed the strain survival for more than two years. Comparing with the strain periodically activated, a critical decrease (about 80%) in -glucan synthesizing capacity was detected for mycelium preserved either on PDY slants or in water. Activation of stored sclerotia followed by subculturing in liquid Production Medium (PM) allowed preparation of homogeneous suspensions for batch fermentations, and scleroglucan concentrations achieved were similar to those with the strain periodically activated.     

Protection of hepatocytes from Fas-mediated apoptosis by a non-transforming SV40 T-antigen mutant

Apoptosis is crucial for the normal development of multicellular organisms and is also important for clearing injured cells, such as virus-infected cells or cancer cells. Defective regulation of apoptosis may contribute to viral pathogenesis and aetiology of cancer. Apoptosis of injured cells is principally triggered by the immune system through cytokines such as Fas-ligand and TNF-alpha. Thus, one of the functions of a viral oncogene, such as SV40T-antigen, may be to inhibit cytokine-mediated apoptosis. We previously demonstrated that Fas-mediated apoptosis of hepatocytes is blocked by the wild-type SV40T-antigen during hepatocarcinogenesis. We determined whether this inhibition was directly related to the T-antigen or whether it is a secondary event of cell transformation, by generating transgenic mice expressing a non-transforming T-antigen mutant able to bind endogenous p53 in the liver. This T-antigen mutant cannot induce hepatocarcinoma, unlike the wild-type T-antigen. However, like the wild-type T-antigen, the mutant was a potent inhibitor of apoptosis induced by the Fas-receptor, but not by the TNF-receptor. Therefore, SV40T-antigen has a new property; the inhibition of Fas-mediated apoptosis, which could facilitate the emergence of transformed hepatocytes, but is not sufficient to induce it.     

Characterization of SKM1, a Saccharomyces cerevisiae gene encoding a novel Ste20/PAK-like protein kinase

Ste20/PAK serine/threonine protein kinases have been suggested as playing essential roles in cell signalling and morphogenesis as potential targets of Cdc42 and Rac GTPases. We have isolated and characterized the Saccharomyces cerevisiae SKM1 gene, which codes for a novel member of this family of protein kinases. The amino acid sequence analysis of Skm1p revealed the presence of a PH domain and a putative p21-binding domain near its amino terminus, suggesting its involvement in cellular signalling or cytoskeletal functions. However, deletion of SKM1 produced no detectable phenotype under standard laboratory conditions. Moreover, disruption of each of the two other S. cerevisiae Ste20/PAK-like kinase-encoding genes, STE20 and CLA4 , in skm1 backgrounds, showed that Skm1p is not redundant with Ste20p or Cla4p. Interestingly, overexpression of SKM1 led to morphological alterations, indicating a possible role for this protein in morphogenetic control. Furthermore, overproduction of Skm1p lacking its N-terminus caused growth arrest. This effect was also seen when similarly truncated versions of Ste20p or Cla4p were overexpressed. We further observed that overproduction of this C-terminal fragment of Skm1p complements the mating defect of a ste20 mutant strain. These results suggest that the N-terminal domains of S. cerevisiae Ste20/PAK-like protein kinases share a negative regulatory function and play a role in substrate specificity.     

Differential effects of five types of antipathogenic plant peptides on model membranes

The effects of five antipathogenic plant peptides, wheat α-thionin, potato PTH1 defensin, barley LTP2 lipid transfer protein, and potato tuber DL1 and DL2 defensins, have been tested against phospholipid vesicles (liposomes). Wheat thionin very actively induces aggregation and leakage of negatively charged vesicles. LTP2 displays the same activities, although to a limited extent. Under certain conditions PTH1 and DL2 induce vesicle aggregation, but not leakage. Potato defensin DL1 failed to show any effect on liposomes. The same peptides have been assayed against a plant pathogenic bacterium, both the membrane-active and -inactive compounds having efficient antibacterial action.     

Analysis of food stimulation of gastrin release in dogs by a panel of inhibitors

The release of gastrin into the serum of five conscious gastric fistula dogs after a meat meal was monitored for 2 hours. Neither the rate of increase in serum gastrin nor the 2 hour cumulative integrated gastrin response was changed by administration of small doses of somatostatin tetradecapeptide (0.5 microgram/kg.hr IV for 2 hr), 16-16 dimethyl prostaglandin E2 (0.25 microgram/kg.hr IV for 2 hr or 1 microgram/kg intragastrically), or bethanechol (20 micrograms/kg.hr IV for 2 hr). Acidification of the food in the antrum to pH 1.2 to 1.4 eliminated serum gastrin release in response to food. In control studies, serum gastrin levels were not altered by IV administration of saline for 2 hr with no food or when a plate of food was held just out of the dogs' reach (teasing). Food-stimulated gastrin release was contrasted with that stimulated by bombesin under identical laboratory conditions [17]. In each case, antral acidification, somatostatin, prostaglandin E2 and bethanechol affected bombesin-stimulated gastrin release differently from that stimulated by food. We conclude that food and bombesin release gastrin by different pathways.     

Studies on plating efficiency and estimation of viability of suspensions of Paracoccidioides brasiliensis yeast cells

Mild sonication was used to obtain single cell suspensions of Paracoccidioides brasiliensis. These cells were intact by microscopic criteria. Direct cell counts in a given inoculum and colony formation on various media were used to determine plating efficiency. Sonicated and nonsonicated cell suspensions were used to study plating efficiency and to estimate viability by means of vital dyes. Methylene blue, Erythrosin B, and Janus green were unreliable when used with P. brasiliensis, but vital dyes were accurate when tested with Candida albicans.Acridine orange gave more meaningful results of viability. Estimates of viability, however, changed significantly as a result of relatively minor alterations in the composition of the suspending medium.In initial experiments, the plating efficiency of P. brasiliensis was dismally low. It descended abruptly with increasing dilution of inoculum. Efficiency was much improved if horse serum was added to brain heart infusion plates or if glucose glycine yeast extract (GGY) plates were incubated at room temperature and mycelial colonies were counted. With the technique we report, current plating efficiency of sonicated suspensions is of the order of 25 %. Our results and procedures have an important bearing upon those studies concerned with in vitro killing of P. brasiliensis in suspensions or with isolating this fungus from clinical or environmental specimens.     

A reliable and sensitive method for the simultaneous determination of dopamine, noradrenaline, 5-hydroxytryptamine and 5-hydroxy-indolacetic acid in small brain samples

A sensitive and reliable fluorometric method for the simultaneous determination of dopamine, noradrenaline, 5-hydroxytryptamine and 5-hydroxy-indol acetic acid in small samples of brain tissues is described. The procedure is based on solvent extraction; catecholamines are oxidized by the Chang's method and 5-hydroxytryptamine and 5-hydroxy-indol acetic acid determined by reaction with o-phthalaldehyde, alpha-methyl-p-tyrosine causes a negligible interference with the procedure. Results of determination of these amines in different brain areas are reported.