Knockouts of Physcomitrella patens CHX1 and CHX2 transporters reveal high complexity of potassium homeostasis.

This study aims to increase our understanding of the functions of CHX transporters in plant cells using the model plant Physcomitrella patens, in which four CHX genes have been identified, PpCHX1-PpCHX4. 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 that they mediated K+ uptake that may be energized by symport with H+. In contrast, these genes suppressed the defect associated with the kha1 mutation in Saccharomyces cerevisiae, which suggests that they might mediate K+/H+ antiport. PpCHX1-green fluorescent protein (GFP) fusion 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 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 slightly higher Rb+ retention than wild-type plants, which suggests that PpCHX2 mediates the transfer of Rb+ either from the vacuole to the cytosol or from the cytosol to the external medium in parallel with other transporters. The distinction between these two possibilities is technically difficult. 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.

(13)C/(12)C isotope labeling to study carbon partitioning and dark respiration in cereals subjected to water stress.

Despite the relevance of carbon (C) loss through respiration processes (with its consequent effect on the lower C availability for grain filling), little attention has been given to this topic. Literature data concerning the role of respiration in cereals are scarce and these have been produced using indirect methods based on gas-exchange estimations. We have developed a new method based on the capture of respired CO(2) samples and their analysis by gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS). In order to analyse the main processes involved in the C balance during grain filling (photosynthesis, respiration, allocation and partitioning) the ambient isotopic (13)C/(12)C composition (delta(13)C) of the growth chamber was modified during this period (delta(13)C ca. -12.8 +/- 0.3 per thousand to ca. -20.0 +/- 0.2 per thousand). The physiological performance, together with the C allocation on total organic matter (TOM) and respiration of wheat (Triticum aestivum L., var. Califa sur) and two hybrids, tritordeum (X Tritordeum Asch. & Graebn line HT 621) and triticale (X Triticosecale Wittmack var. Imperioso), were compared during post-anthesis water stress. In spite of the larger ear DM/total ratio, especially under drought conditions, the grain filling of triticale and wheat was mainly carried out with pre-anthesis C, while the majority of C assimilated during post-anthesis was invested in respiration processes. In the case of wheat and tritordeum, the C balance data suggested a reallocation during grain filling of photoassimilates stored prior to anthesis from shoot to ear. Furthermore, the lower percentage of labeled C on respired CO(2) of droughted tritordeum plants, together with the lower plant biomass, explained the fact that those plants had more C available for grain filling.