Molecular basis of salt tolerance in Physcomitrella Patens model plant: potassium homeostasis and physiological roles of chx transporters

El suelo salino impone un estrés abiótico importante que causa graves problemas en la agricultura ya que la mayoría de los cultivos se ven afectados por la salinidad debido a efectos osmóticos y tóxicos. Por ello, la contaminación y la escasez de agua dulce, la salinización progresiva de tierras y el aumento exponencial de la población humana representan un grave problema que amenaza la seguridad alimentaria mundial para las generaciones futuras. Por lo tanto, aumentar la tolerancia a la salinidad de los cultivos es un objetivo estratégico e ineludible para garantizar el suministro de alimentos en el futuro. Mantener una óptima homeostasis de K+ en plantas que sufren estrés salino es un objetivo importante en el proceso de obtención de plantas tolerantes a la salinidad. Aunque el modelo de la homeostasis de K+ en las plantas está razonablemente bien descrito en términos de entrada de K+, muy poco se sabe acerca de los genes implicados en la salida de K+ o de su liberación desde la vacuola. En este trabajo se pretende aclarar algunos de los mecanismos implicados en la homeostasis de K+ en plantas. Para ello se eligió la briofita Physcomitrella patens, una planta no vascular de estructura simple y de fase haploide dominante que, entre muchas otras cualidades, hacen que sea un modelo ideal. Lo más importante es que no sólo P. patens es muy tolerante a altas concentraciones de Na+, sino que también su posición filogenética en la evolución de las plantas abre la posibilidad de estudiar los cambios claves que, durante el curso de la evolución, se produjeron en las diversas familias de los transportadores de K+. Se han propuesto varios transportadores de cationes como candidatos que podrían tener un papel en la salida de K+ o su liberación desde la vacuola, especialmente miembros de la familia CPA2 que contienen las familias de transportadores KEA y CHX. En este estudio se intenta aumentar nuestra comprensión de las funciones de los transportadores de CHX en las células de las plantas usando P. patens, como ya se ha dicho. En esta especie, se han identificado cuatro genes CHX, PpCHX1-4. Dos de estos genes, PpCHX1 y PpCHX2, se expresan aproximadamente al mismo nivel que el gen PpACT5, y los otros dos genes muestran una expresión muy baja. La expresión de PpCHX1 y PpCHX2 en mutantes de Escherichia coli defectivos en el transporte de K+ restauraron el crecimiento de esta cepa en medios con bajo contenido de K+, lo que viii sugiere que la entrada de K+ es energizada por un mecanismo de simporte con H+. Por otra parte, estos transportadores suprimieron el defecto asociado a la mutación kha1 en Saccharomyces cerevisiae, lo que sugiere que podrían mediar un antiporte en K+/H+. La proteína PpCHX1-GFP expresada transitoriamente en protoplastos de P. patens co-localizó con un marcador de Golgi. En experimentos similares, la proteína PpCHX2-GFP localizó aparentemente en la membrana plasmática y tonoplasto. Se construyeron las líneas mutantes simples de P. patens ΔPpchx1 y ΔPpchx2, y también el mutante doble ΔPpchx2 ΔPphak1. Los mutantes simples crecieron normalmente en todas las condiciones ensayadas y mostraron flujos de entrada normales de K+ y Rb+; la mutación ΔPpchx2 no aumentó el defecto de las plantas ΔPphak1. En experimentos a largo plazo, las plantas ΔPpchx2 mostraron una retención de Rb+ ligeramente superior que las plantas silvestres, lo que sugiere que PpCHX2 promueve la transferencia de Rb+ desde la vacuola al citosol o desde el citosol al medio externo, actuando en paralelo con otros transportadores. Sugerimos que transportadores de K+ de varias familias están involucrados en la homeostasis de pH de orgánulos ya sea mediante antiporte K+/H+ o simporte K+-H+.ix ABSTRACT Soil salinity is a major abiotic stress causing serious problems in agriculture as most crops are affected by it. Moreover, the contamination and shortage of freshwater, progressive land salinization and exponential increase of human population aggravates the problem implying that world food security may not be ensured for the next generations. Thus, a strategic and an unavoidable goal would be increasing salinity tolerance of plant crops to secure future food supply. Maintaining an optimum K+ homeostasis in plants under salinity stress is an important trait to pursue in the process of engineering salt tolerant plants. Although the model of K+ homeostasis in plants is reasonably well described in terms of K+ influx, very little is known about the genes implicated in K+ efflux or release from the vacuole. In this work, we aim to clarify some of the mechanisms involved in K+ homeostasis in plants. For this purpose, we chose the bryophyte plant Physcomitrella patens, a nonvascular plant of simple structure and dominant haploid phase that, among many other characteristics, makes it an ideal model. Most importantly, not only P. patens is very tolerant to high concentrations of Na+, but also its phylogenetic position in land plant evolution opens the possibility to study the key changes that occurred in K+ transporter families during the course of evolution. Several cation transporter candidates have been proposed to have a role in K+ efflux or release from the vacuole especially members of the CPA2 family which contains the KEA and CHX transporter families. We intended in this study to increase our understanding of the functions of CHX transporters in plant cells using P. patens, in which four CHX genes have been identified, PpCHX1-4. Two of these genes, PpCHX1 and PpCHX2, are expressed at approximately the same level as the PpACT5 gene, but the other two genes show an extremely low expression. PpCHX1 and PpCHX2 restored growth of Escherichia coli mutants on low K+-containing media, suggesting they mediated K+ uptake that may be energized by symport with H+. In contrast, these genes suppressed the defect associated to the kha1 mutation in Saccharomyces cerevisiae, which suggest that they might mediate K+/H+ antiport. PpCHX1-GFP protein transiently expressed in P. patens protoplasts co-localized with a Golgi marker. In similar experiments, the PpCHX2-GFP protein appeared to localize to tonoplast and plasma x membrane. We constructed the ΔPpchx1 and ΔPpchx2 single mutant lines, and the ΔPpchx2 ΔPphak1 double mutant. Single mutant plants grew normally under all the conditions tested and exhibited normal K+ and Rb+ influxes; the ΔPpchx2 mutation did not increase the defect of ΔPphak1 plants. In long-term experiments, ΔPpchx2 plants showed a slightly higher Rb+ retention than wild type plants, which suggests that PpCHX2 mediates the transfer of Rb+ from either the vacuole to the cytosol or from the cytosol to the external medium in parallel with other transporters. We suggest that K+ transporters of several families are involved in the pH homeostasis of organelles by mediating either K+/H+ antiport or K+-H+ symport.

Geochemistry of Baltic Sea sediments and porewaters

Patterns of regeneration and burial of phosphorus (P) in the Baltic Sea are strongly dependent on redox conditions. Redox varies spatially along water depth gradients and temporally in response to the seasonal cycle and multidecadal hydrographic variability. Alongside the well-documented link between iron oxyhydroxide dissolution and release of P from Baltic Sea sediments, we show that preferential remineralization of P with respect to carbon (C) and nitrogen (N) during degradation of organic matter plays a key role in determining the surplus of bioavailable P in the water column. Preferential remineralization of P takes place both in the water column and upper sediments and its rate is shown to be redox-dependent, increasing as reducing conditions become more severe at greater water-depth in the deep basins. Existing Redfield-based biogeochemical models of the Baltic may therefore underestimate the imbalance between N and P availability for primary production, and hence the vulnerability of the Baltic to sustained eutrophication via the fixation of atmospheric N. However, burial of organic P is also shown to increase during multidecadal intervals of expanded hypoxia, due to higher net burial rates of organic matter around the margins of the deep basins. Such intervals may be characterized by basin-scale acceleration of all fluxes within the P cycle, including productivity, regeneration and burial, sustained by the relative accessibility of the water column P pool beneath a shallow halocline.

(Table 2B) Geochemistry of Ariadne II sediments corrected for dilution by salt from pore waters and are carbonate free in the eastern South Pacific

Sediment depth is given in mbsf.

Physical and chemical properties, 14C ages and palynology of sediment cores from the Western Baltic Sea

Sea level related radiocarbon, palynological and stratigraphical data from sediment cores in the Western Baltic have been tested against the existing sea level curves for the region. The relative sea level rise curves for the beginning of the Holocene show no significant deviations between the Kiel, Mecklenburg und Lübeck Bays and hence do not support the previously reported differences in the averaged regional subsidence rates for this time interval. Local subsidence and upheaval due to salt tectonics probably played a greater role than previously suspected in the region. The sea level possibly stagnated around -28 m during the early Holocene before rising very rapidly to -14 m. The submarine terraces at -30 m and perhaps also at -27 m were formed during the lacustrine phase of the Western Baltic when the water levels were controlled by the main thresholds in the Great Belt.

Sedimentology of the southern Bellingshausen Sea

A major trough ('Belgica Trough') eroded by a palaeo-ice stream crosses the continental shelf of the southern Bellingshausen Sea (West Antarctica) and is associated with a trough mouth fan ('Belgica TMF') on the adjacent continental slope. Previous marine geophysical and geological studies investigated the bathymetry and geomorphology of Belgica Trough and Belgica TMF, erosional and depositional processes associated with bedform formation, and the temporal and spatial changes in clay mineral provenance of subglacial and glaciomarine sediments. Here, we present multi-proxy data from sediment cores recovered from the shelf and uppermost slope in the southern Bellingshausen Sea and reconstruct the ice-sheet history since the last glacial maximum (LGM) in this poorly studied area of West Antarctica. We combined new data (physical properties, sedimentary structures, geochemical and grain-size data) with published data (shear strength, clay mineral assemblages) to refine a previous facies classification for the sediments. The multi-proxy approach allowed us to distinguish four main facies types and to assign them to the following depositional settings: 1) subglacial, 2) proximal grounding-line, 3) distal sub-ice shelf/subsea ice, and 4) seasonal open-marine. In the seasonal open-marine facies we found evidence for episodic current-induced winnowing of near-seabed sediments on the middle to outer shelf and at the uppermost slope during the late Holocene. In addition, we obtained data on excess 210Pb activity at three core sites and 44 AMS 14C dates from the acid-insoluble fraction of organic matter (AIO) and calcareous (micro-)fossils, respectively, at 12 sites. These chronological data enabled us to reconstruct, for the first time, the timing of the last advance and retreat of the West Antarctic Ice Sheet (WAIS) and the Antarctic Peninsula Ice Sheet (APIS) in the southern Bellingshausen Sea. We used the down-core variability in sediment provenance inferred from clay mineral changes to identify the most reliable AIO 14C ages for ice-sheet retreat. The palaeo-ice stream advanced through Belgica Trough after ~36.0 corrected 14C ka before present (B.P.). It retreated from the outer shelf at ~25.5 ka B.P., the middle shelf at ~19.8 ka B.P., the inner shelf in Eltanin Bay at ~12.3 ka B.P., and the inner shelf in Ronne Entrance at ~6.3 ka B.P.. The retreat of the WAIS and APIS occurred slowly and stepwise, and may still be in progress. This dynamical ice-sheet behaviour has to be taken into account for the interpretation of recent and the prediction of future mass-balance changes in the study area. The glacial history of the southern Bellingshausen Sea is unique when compared to other regions in West Antarctica, but some open questions regarding its chronology need to be addressed by future work.

Propagation of Spartina alterniflora for substrate stabilization and salt marsh development /

"August 1974."

Response of the salt-freshwater interface in a coastal aquifer to a wave-induced groundwater pulse: field observations and modelling

Field observations on an unconfined coastal aquifer showed that a groundwater pulse, generated by it moderate (significant wave height, H-sig similar to 4.5 m) wave/storm event, induced significant oscillations in the salt-freshwater interface of the order of several metres in the horizontal direction. A dynamic sharp-interface model is developed to quantify the mechanism of these interface oscillations. The model uses the 50% seawater salinity contour as the location of the equivalent sharp-interface. The model was calibrated against the observed groundwater table fluctuations. It predicted reasonably well the interface oscillations with a slight over-prediction of the oscillation magnitude and a steepening of the interface. The neglect of mixing in the salt-freshwater mixing zone by the sharp-interface model is suggested as a possible contributor to the discrepancies between the model predictions and observations. In contrast with the significant wave effects, there was no observable response of the interface to diurnal or semidiurnal tides. (C) 2004 Elsevier Ltd. All rights reserved.

Cholinergic and adrenergic innervation of lingual salt glands of the estuarine crocodile, Crocodylus porosus

Many marine reptiles and birds possess extrarenal salt glands that facilitate the excretion of excess sodium and chloride ions accumulated as a consequence of living in saline environments. Control of the secretory activity of avian salt glands is under neural control, but little information is available on the control of reptilian salt glands. Innervation of the lingual salt glands of the salt water crocodile, Crocodylus porosus, was examined in salt water-acclimated animals using histological methods. Extensive networks of both cholinergic and adrenergic nerve fibres were identified close to salt-secreting lobules and vasculature. The identification of both catecholamine-containing and cholinergic neurons in the salt gland epithelium and close to major blood vessels in the tissue suggests the action of the neurotransmitters on the salt-secreting epithelium itself and the rich vascular network of the lingual salt glands.