Structural features of an arabinogalactan gum exudates from Spondias dulsis (Anacardiaceae)

The tree Spondias dulcis, located in Venezuela, exudes a light-brown gum. The polysaccharide, isolated from the original gum, contains galactose, arabinose, mannose, rhamnose, glucuronic acid, and its 4-O-methyl derivative. Application of chemical methods, in combination with 1D and 2D NMR spectroscopy afforded interesting structural features of the gum polysaccharide. The unequivocal presence of rhamnose in the polymer structure was confirmed by chemical and spectral data [1H (1.03 ppm); 13C (16.92 ppm)]. Also confirmed was the existence of 3-O- and 6-O-substitutes galactose residues by the spectral data correlations observed in Heteronuclear Multiple Quantum Coherence (HMQC) and Heteronuclear Multiple Bond Correlation (HMBC). Also observed were unequivocal resonances for beta-D-glucuronic acid and its 4-O-methyl derivative, and the presence of 3-O-alpha-L-arabinofuranose and 3-O-beta-L-arabinopyranose residues.     

Genetic structure of populations of Calomys laucha (muridae, sigmodontinae) from central Argentina

The genetic structure of populations of the Sigmodontinae rodent Calomys laucha was studied by means of allozyme electrophoresis. This highly opportunistic species is found preferably in periodically perturbed habitats of crop fields in central Argentina, where it can attain very high densities. A total of 17 enzymatic proteins assayed gave information on 25 loci; only four were monomorphic in the seven populations studied. Levels of genetic variability (H_o from 0.144 to 0.171; P_95% from 44% to 56%) were higher than mean values reported for mammals and rodents. These high levels of heterozygosity could be maintained by large populations that do not experience great fluctuations in size, or by a social structure consistent in many small breeding groups that are formed and dissappear every breeding season. Genetic differentiation at a macrogeographical scale (θ=0.018) was low but statistically significant, and showed no correlation with geographic distance between pairs of populations. The pattern of population differentiation found is compatible with a relatively recent range expansion.     

Oligoryzomys flavescens (Rodentia, Muridae): gene flow among populations from central-eastern Argentina

In species acting as hosts of infectious agents, the extent of gene flow between populations is of particular interest because the expansion of different infectious diseases is usually related to the dispersal of the host. We have estimated levels of gene flow among populations of the sigmodontine rodent Oligoryzomys flavescens, in which high titers of antibodies have been detected for a Hantavirus in Argentina that produces a severe pulmonary syndrome. Enzyme polymorphism was studied by means of starch gel electrophoresis in 10 populations from the area where human cases of Hantavirus have occurred. Genetic differentiation between populations was calculated from F_ST values with the equation Nm = [(1/F_ST−1]/4. To assess the relative importance of current gene flow and historical associations between populations, the relationship of population pairwise log Nm and log geographic distance was examined. Low F_ST (mean = 0.038) and high Nm (15.27) values suggest high levels of gene flow among populations. The lack of an isolation by distance pattern would indicate that this species has recently colonized the area. The northernmost population, located on the margin of a great river, shows very high levels of gene flow with the downstream populations despite the large geographic distances. Passive transport of animals down the river by floating plants would promote unidirectional gene flow. This fact and the highest mean heterozygosity of that northernmost population suggest it is a center of dispersal within the species' range. This revised version was published online in July 2006 with corrections to the Cover Date.     

Structures of the signal recognition particle receptor from the archaeon Pyrococcus furiosus: implications for the targeting step at the membrane

In all organisms, a ribonucleoprotein called the signal recognition particle (SRP) and its receptor (SR) target nascent proteins from the ribosome to the translocon for secretion or membrane insertion. We present the first X-ray structures of an archeal FtsY, the receptor from the hyper-thermophile Pyrococcus furiosus (Pfu), in its free and GDP*magnesium-bound forms. The highly charged N-terminal domain of Pfu-FtsY is distinguished by a long N-terminal helix. The basic charges on the surface of this helix are likely to regulate interactions at the membrane. A peripheral GDP bound near a regulatory motif could indicate a site of interaction between the receptor and ribosomal or SRP RNAs. Small angle X-ray scattering and analytical ultracentrifugation indicate that the crystal structure of Pfu-FtsY correlates well with the average conformation in solution. Based on previous structures of two sub-complexes, we propose a model of the core of archeal and eukaryotic SRP*SR targeting complexes.     

Predator feeding ecology on Patagonian rocky outcrops: implications for colonies of mountain vizcacha (Lagidium viscacia)

On 10 rocky outcrops with mountain vizcacha colonies, diets and trophic relations of five predator species were analyzed to evaluate their impact in vizcacha populations. Diets included 17 prey items of which mammals represented 97% of biomass. Lagomorphs were the most important prey, followed by sigmodontines, tuco-tucos, and mountain vizcachas. The Barn Owl was a specialized predator, while the culpeo and lesser grison were generalized consumers. Predation of mountain vizcacha by Magellanic Horned Owls and culpeo foxes could increase vizcacha vulnerability because vizcacha populations are small and fragmented, and females produce only one offspring per year.     

VopA inhibits ATP binding by acetylating the catalytic loop of MAPK kinases

The bacterial pathogen Vibrio parahemeolyticus manipulates host signaling pathways during infections by injecting type III effectors. One of these effectors, Vibrio outer protein A (VopA), inhibits MAPK signaling via a novel mechanism, distinct from those described for other bacterial toxins, that disrupts this signaling pathway. VopA is an acetyltransferase that potently inhibits MAPK signaling pathways not only by preventing the activation of MAPK kinases (MKKs) but also by inhibiting the activity of activated MKKs. VopA acetylates a conserved lysine found in the catalytic loop of all kinases and blocks the binding of ATP, but not ADP, on the MKKs, resulting in an inactive phosphorylated kinase. Acetylation of this conserved lysine inhibits kinase activity by a new mechanism of regulation that has not been observed previously. Identifying the target of VopA reveals a way that the reversible post-translational modification of lysine acetylation can be used to regulate the activity of an enzyme.     

A Rapid and Reliable Method for Total Protein Extraction from Succulent Plants for Proteomic Analysis

Crassulacean acid metabolism plants have some morphological features, such as succulent and reduced leaves, thick cuticles, and sunken stomata that help them prevent excessive water loss and irradiation. As molecular constituents of these morphological adaptations to xeric environments, succulent plants produce a set of specific compounds such as complex polysaccharides, pigments, waxes, and terpenoids, to name a few, in addition to uncharacterized proteases. Since all these compounds interfere with the analysis of proteins by electrophoretic techniques, preparation of high quality samples from these sources represents a real challenge. The absence of adequate protocols for protein extraction has restrained the study of this class of plants at the molecular level. Here, we present a rapid and reliable protocol that could be accomplished in 1 h and applied to a broad range of plants with reproducible results. We were able to obtain well-resolved SDS/PAGE protein patterns in extracts from different members of the subfamilies Agavoideae (Agave, Yucca, Manfreda, and Furcraea), Nolinoideae (Dasylirion and Beucarnea), and the Cactaceae family. This method is based on the differential solubility of contaminants and proteins in the presence of acetone and pH-altered solutions. We speculate about the role of saponins and high molecular weight carbohydrates to produce electrophoretic-compatible samples. A modification of the basic protocol allowed the analysis of samples by bidimensional electrophoresis (2DE) for proteomic analysis. Furostanol glycoside 26-O-β-glucosidase (an enzyme involved in steroid saponin synthesis) was successfully identified by mass spectrometry analysis and de novo sequencing of a 2DE spot from an Agave attenuata sample.     

How amyloplasts,water deficit and root tropisms interact?

Hydrotropism, the differential growth of plant roots directed by a moisture gradient, is a long recognized, but not well-understood plant behavior. Hydrotropism has been characterized in the model plant Arabidopsis. Previously, it was postulated that roots subjected to water stress are capable of undergo water-directed tropic growth independent of the gravity vector because of the loss of the starch granules in root cap columella cells and hence the loss of the early steps in gravitropic signaling. We have recently proposed that starch degradation in these cells during hydrostimulation sustain osmotic stress and root growth for carrying out hydrotropism instead of reducing gravity responsiveness. In addition, we also proposed that abscisic acid (ABA) and water deficit are critical regulators of root gravitropism and hydrotropism, and thus mediate the interacting mechanism between these two tropisms. Our conclusions are based upon experiments performed with the no hydrotropic response (nhr1) mutant of Arabidopsis, which lacks a hydrotropic response and shows a stronger gravitropic response than that of wild type (WT) in a medium with an osmotic gradient.Key words: starch, water deficit, auxin, abscisic acid, gravitropism, hydrotropismRoots of land plants sense and respond to different stimuli, some of which are fixed in direction and intensity (i.e., gravity) while other vary in time, space, direction and intensity (i.e., obstacles and moisture gradients). Directed growth of roots in relation to a gradient in moisture is called hydrotropism and begins in the root cap with the sensing of the moisture gradient. However, since gravity is an omnipresent accompaniment of Earthly life and many living process have evolved with it as a background constant, it is not surprising that root hydrotropism interacts with gravitropism.¹ The hydrotropic response in Arabidopsis, compare with other plants such as pea and cucumber^2,3 is readily observed even in the presence of gravity.^4,5 When Arabidopsis roots are subjected to a water gradient, such that the source of water is placed 180° opposed to the gravity vector, the roots will grow upwards, displaying positive hydrotropism. Therefore, it has been feasible to isolate so far two Arabidopsis mutants affected in their hydrotropic response.^5,6 Analysis of these mutants reveals new insights of the mechanism of hydrotropism. For one hand, the no hydrotropic response (nhr1) mutant lacks a hydrotropic response, and shows a stronger gravitropic response than that of wt and a modified wavy growth response in a medium with an osmotic gradient.^5,7 On the other hand, the mizu-kussei1 (miz1) mutant did not exhibit hydrotropism and showed regular gravitropism.⁶ Hence, the root hydrotropic response is both linked and unlinked from the gravitropic one. Nonetheless, miz1 roots also showed a reduced phototropism and a modified wavy growth response. This indicates that both MIZ1 and NHR1 are not exclusive components of the mechanism for hydrotropism and supports the notion that the root cap has assessment mechanisms that integrate many different environmental influences to produce a final integrated response.⁸ Thus, the physiological phenomena distinctively displayed by roots in order to forage resources from the environment are the result of integrated responses that resulted from many environmental influences sensed in the root cap.In the course of studying how gravity and water availability affected the perception and assessment of each other in root cap cells that generated the final root tropic response, we found that ABA is a critical regulator of the signal transduction mechanism that integrated these two-root tropisms.⁷ For this, we analyzed the long-term hydrotropic response of Arabidopsis roots in an osmotic gradient system. ABA, locally applied to seeds or root tips of nhr1, significantly increased root downward growth in a medium with an osmotic gradient (root length of nhr1 seedlings grown in this medium were on average 12.5 mm and plus 10 µM ABA were 25.1 mm). On the other hand, WT roots germinated and treated locally with ABA in this system were strongly gravitropic, albeit they had almost no starch in amyloplasts of root cap columella cells. Hydrotropically stimulated nhr1 roots, with or without ABA, maintained starch in amyloplastas, as opposed to those of WT. Therefore, the near-absence (WT) or abundant presence (nhr1) of starch granules does not affect the extent of downward gravitropism of roots in an osmotic gradient medium. Starch degradation in the wt might participate in osmoregulation by which root cells maintain turgor and consequently carry out hydrotropism, instead of reducing gravity responsiveness. In fact, it was just recently published that salt-induced rapid degradation of starch in amyloplasts is not likely the main reason for a negative gravitropic response seen under salt stress, because sos mutant roots of Arabidopsis showed negative gravitropic growth without any apparent rapid digestion of starch granules.⁹ Additionally, the stems of overwintering tubers of Potamogeton pectinatus are capable of elongating much faster in the absence than in the presence of oxygen for up to 14 days and its stems has an enhanced capacity for gravitropic movements in completely anoxic conditions.¹⁰ These authors hypothesized that ABA and starch degradation in the starchy tuber sustained stem cell elongation and cell division as well as differential growth required for the gravitropic response in these aquatic plants. These data taken together suggest that in conditions of anoxia, or water stress, ABA and degradation of starch play a critical role in the ability to survive relatively prolonged periods of unfavorable growth conditions. These players are critical when water or minerals are scarce since they regulate the enhancement of root downward growth. However, since roots can trail humidity gradients in soil, they can modulate their branching patterns (architecture) and thus respond to hydrotropism once a water-rich patch is found. Then the response of plants to gravity is principally one of nutrition (shoots to light, roots to mineral and water) and consequently must be regulated according to the long- and short-term environmental variables that occur during the development of the plant.Differential growth that occurs during the gravitropic and phototropic response has been explained according to the Cholodny-Went hypothesis, which states that the lateral transport of auxin across stimulated plant tissues is responsible for the curvature response.¹¹ Analysis of hydrotropism in some Arabidopsis agravitropic auxin transport mutants has demonstrated that these mutations do not influence their hydrotropic response.⁴ Furthermore, current pharmacological studies using inhibitors also indicated that both auxin influx and efflux are not required for hydrotropic response whereas auxin response is necessary for it.¹² These authors suggested a novel mechanism for auxin in root hydrotropism. Here, we analyzed whether asymmetric auxin distribution takes place across hydrotropically-stimulated roots using transgenic plants carrying a responsive auxin promoter (DR5) driving the expression of β-glucuronidase (GUS) or green fluorescent protein (GFP)^13,14 in wt and nhr1 backgrounds. Wt and nhr1 roots hydrotropically stimulated in a system with air moisture gradient⁵ showed no asymmetric expression of the DR5:: GUS or DR5::GFP (Fig. 1A and B). Nonetheless, nhr1 roots showed a substantial decrease in the signal driven by the DR5::GUS and GFP reporters in humidity saturated conditions (Fig. 1A, part b and B, part b), which might indicate that auxin-induced gene expression in the root cap was inhibited. It remains to be determined the significance of this inhibition in the no hydrotropic response phenotype displayed by nhr1 roots. Determination of the DR5::GUS expression in wt and nhr1 roots growing in an osmotic gradient medium for testing long-term hydrotropism revealed that the GUS signal was to some extent diminished in both wt or in nhr1 roots (Fig. 2C and D) compared to those roots growing in normal medium (Fig. 2A and B). An inhibitor of auxin response reduced hydrotropism,¹² and also inhibited auxin-dependent DR5::GUS expression.¹⁵ However, a decrease of DR5::GUS in wt root tips was not an impediment for developing an hydrotropic response. On the other hand, nhr1 roots also showed a decrease of DR5::GUS expression (Fig. 2B and D) and a complete absence of DR5::GFP (data not shown), which did not influence the extent of downward root gravitropism in water deficit conditions. Therefore, it is difficult to assign a role of auxin-induce gene expression in hydrotropism and further studies are required in order to unravel this issue. Furthermore, it needs to be resolved whether these expression studies oppose the idea that gradients in auxin precede differential growth in response to humidity gradients.Open in a separate windowFigure 1DR5:: GUS (A) and DR5::GFP (B) activity in the wild type NHR1 and nhr1 backgrounds. (A) Root tips hydrostimulated in a system with air moisture gradient (C and D) or grown in a saturated water conditions (A and B) stained with 1 mM 5-bromo-4-chloro-3-indolyl-β-d-glucuronic (X-Gluc) acid buffer under the same conditions for 80 min. (B) Root tips hydrostimulated as in (A) (C and D) or grown in a saturated water conditions (A and B) whose green fluorescent signal was visualized by confocal microscopy. Shown are images selected from at least 45 representative root tips. Bar = 29 µm.Open in a separate windowFigure 2Expression of DR5::GUS in wild type NHR1 and nhr1 backgrounds. Roots were hydrotropically stimulated for 8 days in a medium with an osmotic gradient (C and D) or grown in normal medium (A and B) and stained with X-Gluc acid buffer under the same conditions for 80 min. Shown are images selected from at least 50 representative root tips. Bar = 25 µm.Our studies⁷ revealed that ABA is a critical regulator of both root gravitropism and hydrotropism in water deficit conditions, and that the role of auxin under these conditions seems to differ from those observed in several studies thus far published on gravitropism made under well-water conditions. The molecular characterization of NHR1 and from other nhr-like mutants already isolated in our lab will clarify the mechanisms involved in this fascinating tropism.¹⁶     

Temporal dynamics and genetic diversity of chemotactic‐competent microbial populations in the rhizosphere

The contribution of chemotaxis to the competitive colonization of the rhizosphere for the vast majority of the soil community is unknown. We have developed and applied a molecular diagnostic tool, based on a gene encoding the central regulator of bacterial chemotaxis (cheA), to characterize and temporally track specific populations of native microbes with chemotaxis potential that are present in soil exposed to two rhizospheres: wheat and cowpea. The data show that the chemotactic‐competent communities present in the rhizospheres of the two plants are distinct and less diverse than the bulk soil, indicating the development of unique microbial communities. Consistent with the supposition that selection and recruitment of specific soil microbes takes place in the rhizosphere, the dynamics of specific cheA phylotypes provides support for the hypothesis that chemotaxis provides a competitive advantage to some soil microbes. This is the first study to examine and profile the genetic diversity of chemotaxis genes in natural populations. As such, it illustrates our limited understanding of microbial chemotaxis for the majority of soil microbes. It also highlights the value of a culture‐independent approach for examining chemotaxis populations in order to build empirical lines of evidence for its role in structuring of microbial assemblages.