Phenotype of Arabidopsis thaliana semi-dwarfs with deep roots and high growth rates under water-limiting conditions is independent of the GA5 loss-of-function alleles.

BACKGROUND AND AIMS: The occurrence of Arabidopsis thaliana semi-dwarf accessions carrying inactive alleles at the gibberellin (GA) biosynthesis GA5 locus has raised the question whether there are pleiotropic effects on other traits at the root level, such as rooting depth. In addition, it is unknown whether semi-dwarfism in arabidopsis confers a growth advantage under water-limiting conditions compared with wild-type plants. The aim of this research was therefore to investigate whether semi-dwarfism has a pleiotropic effect in the root system and also whether semi-dwarfs might be more tolerant of water-limiting conditions. METHODS: The root systems of different arabidopsis semi-dwarfs and GA biosynthesis mutants were phenotyped in vitro using the GROWSCREEN-ROOT image-based software. Semi-dwarfs were phenotyped together with tall, near-related accessions. In addition, root phenotypes were investigated in soil-filled rhizotrons. Rosette growth trajectories were analysed with the GROWSCREEN-FLUORO setup based on non-invasive imaging. KEY RESULTS: Mutations in the early steps of the GA biosynthesis pathway led to a reduction in shoot as well as root size. Depending on the genetic background, mutations at the GA5 locus yielded phenotypes characterized by decreased root length in comparison with related wild-type ones. The semi-dwarf accession Pak-3 showed the deepest root system both in vitro and in soil cultivation experiments; this comparatively deep root system, however, was independent of the ga5 loss-of-function allele, as shown by co-segregation analysis. When the accessions were grown under water-limiting conditions, semi-dwarf accessions with high growth rates were identified. CONCLUSIONS: The observed diversity in root system growth and architecture occurs independently of semi-dwarf phenotypes, and is probably linked to a genetic background effect. The results show that there are no clear advantages of semi-dwarfism at low water availability in arabidopsis.

Effects of altered α- and ß-branch carotenoid biosynthesis on photoprotection and whole-plant acclimation of Arabidopsis to photo-oxidative stress.

Functions of α- and ß-branch carotenoids in whole-plant acclimation to photo-oxidative stress were studied in Arabidopsis thaliana wild-type (wt) and carotenoid mutants, lutein deficient (lut2, lut5), non-photochemical quenching1 (npq1) and suppressor of zeaxanthin-less1 (szl1) npq1 double mutant. Photo-oxidative stress was applied by exposing plants to sunflecks. The sunflecks caused reduction of chlorophyll content in all plants, but more severely in those having high α- to ß-branch carotenoid composition (α/ß-ratio) (lut5, szl1npq1). While this did not alter carotenoid composition in wt or lut2, which accumulates only ß-branch carotenoids, increased xanthophyll levels were found in the mutants with high α/ß-ratios (lut5, szl1npq1) or without xanthophyll-cycle operation (npq1, szl1npq1). The PsbS protein content increased in all sunfleck plants but lut2. These changes were accompanied by no change (npq1, szl1npq1) or enhanced capacity (wt, lut5) of NPQ. Leaf mass per area increased in lut2, but decreased in wt and lut5 that showed increased NPQ. The sunflecks decelerated primary root growth in wt and npq1 having normal α/ß-ratios, but suppressed lateral root formation in lut5 and szl1npq1 having high α/ß-ratios. The results highlight the importance of proper regulation of the α- and ß-branch carotenoid pathways for whole-plant acclimation, not only leaf photoprotection, under photo-oxidative stress.

The platform GrowScreen-Agar enables identification of phenotypic diversity in root and shoot growth traits of agar grown plants.

BACKGROUND: Root system architecture and especially its plasticity in acclimation to variable environments play a crucial role in the ability of plants to explore and acquire efficiently soil resources and ensure plant productivity. Non-destructive measurement methods are indispensable to quantify dynamic growth traits. For closing the phenotyping gap, we have developed an automated phenotyping platform, GrowScreen-Agar, for non-destructive characterization of root and shoot traits of plants grown in transparent agar medium. RESULTS: The phenotyping system is capable to phenotype root systems and correlate them to whole plant development of up to 280 Arabidopsis plants within 15 min. The potential of the platform has been demonstrated by quantifying phenotypic differences within 78 Arabidopsis accessions from the 1001 genomes project. The chosen concept 'plant-to-sensor' is based on transporting plants to the imaging position, which allows for flexible experimental size and design. As transporting causes mechanical vibrations of plants, we have validated that daily imaging, and consequently, moving plants has negligible influence on plant development. Plants are cultivated in square Petri dishes modified to allow the shoot to grow in the ambient air while the roots grow inside the Petri dish filled with agar. Because it is common practice in the scientific community to grow Arabidopsis plants completely enclosed in Petri dishes, we compared development of plants that had the shoot inside with that of plants that had the shoot outside the plate. Roots of plants grown completely inside the Petri dish grew 58% slower, produced a 1.8 times higher lateral root density and showed an etiolated shoot whereas plants whose shoot grew outside the plate formed a rosette. In addition, the setup with the shoot growing outside the plate offers the unique option to accurately measure both, leaf and root traits, non-destructively, and treat roots and shoots separately. CONCLUSIONS: Because the GrowScreen-Agar system can be moved from one growth chamber to another, plants can be phenotyped under a wide range of environmental conditions including future climate scenarios. In combination with a measurement throughput enabling phenotyping a large set of mutants or accessions, the platform will contribute to the identification of key genes.

Medicinal plants: a natural chaperones source for treating neurological disorders.

Currently, no pharmaceuticals for the etiological treatment of neurodegenerative protein-misfolding diseases (e.g., ALS, Alzheimer's or prion diseases) are available. In this brief communication the development of chaperone-based medications from medicinal plants (e.g., Ginkgo biloba) are reviewed as referred to specific protein-protein interactions of plant metallochaperones and human enzymes. It is indicated that bioactive copper chaperones for superoxide dismutase isolated from medicinal plants may be lead molecules for drug development in several diseases concerning metal ion metabolisms of man and animals.

New hope for the diagnosis and therapy of Alzheimer's disease.

Improperly folded metal cofactor-containing proteins (e.g., copper chaperone for superoxide dismutase, CCS) are believed to play a key role in several protein-misfolding diseases (e.g., Alzheimer's disease or Amyotrophic Lateral Sclerosis) because under regular physiological conditions, metallochaperones activate or stabilize the native conformation of important metalloproteins (e.g., superoxide dismutase) in certain cellular processes. For an improved diagnosis and therapy of neurodegenerative diseases, new methodologies have to be developed that enable a well-defined differentiation between properly folded and inactive metalloproteins in clinical samples. In the literature it is reported that different high molecular mass metal-containing proteins were isolated in brain samples from Alzheimer's patients and in vegetables by using a 2-dimensional polyacrylamide gel electrophoresis (2-DE) procedure. In the present article, selected results of these studies are scrutinized and compared with some results obtained by a standardized method termed 'quantitative preparative native continuous polyacrylamide gel electrophoresis (QPNC-PAGE)'. Conclusively, QPNC-PAGE is a highly efficient approach used by biochemists to resolve native and denatured metalloproteins (MW 6 - > or = 200 kDa) in complex protein mixtures.

Important contributions of a new quantitative preparative native continuous polyacrylamide gel electrophoresis (QPNC-PAGE) procedure for elucidating metal cofactor metabolisms in protein-misfolding diseases--a theory.

The quantitative analysis of metallochaperone proteins in biofluids (e.g. blood, liquor) may be a major prerequisite for clinical investigations concerning the structure-function relationships of biologically-active metal cofactor-containing chaperones in protein-misfolding diseases (e.g. Alzheimer's or related diseases). For these purposes, a new state-of-the-art gel electrophoresis [quantitative preparative native continuous polyacrylamide gel electrophoresis procedure (QPNC-PAGE)] combined with biological mass and NMR spectrometries might essentially contribute to provide fundamental insights into the metabolisms of important metal cofactors in biological systems and the proper folding of metallochaperones in conformational diseases.

GROWSCREEN-Rhizo is a novel phenotyping robot enabling simultaneous measurements of root and shoot growth for plants grown in soil-filled rhizotrons.

Root systems play an essential role in ensuring plant productivity. Experiments conducted in controlled environments and simulation models suggest that root geometry and responses of root architecture to environmental factors should be studied as a priority. However, compared with aboveground plant organs, roots are not easily accessible by non-invasive analyses and field research is still based almost completely on manual, destructive methods. Contributing to reducing the gap between laboratory and field experiments, we present a novel phenotyping system (GROWSCREEN-Rhizo), which is capable of automatically imaging roots and shoots of plants grown in soil-filled rhizotrons (up to a volume of ~18L) with a throughput of 60 rhizotrons per hour. Analysis of plants grown in this setup is restricted to a certain plant size (up to a shoot height of 80cm and root-system depth of 90cm). We performed validation experiments using six different species and for barley and maize, we studied the effect of moderate soil compaction, which is a relevant factor in the field. First, we found that the portion of root systems that is visible through the rhizotrons' transparent plate is representative of the total root system. The percentage of visible roots decreases with increasing average root diameter of the plant species studied and depends, to some extent, on environmental conditions. Second, we could measure relatively minor changes in root-system architecture induced by a moderate increase in soil compaction. Taken together, these findings demonstrate the good potential of this methodology to characterise root geometry and temporal growth responses with relatively high spatial accuracy and resolution for both monocotyledonous and dicotyledonous species. Our prototype will allow the design of high-throughput screening methodologies simulating environmental scenarios that are relevant in the field and will support breeding efforts towards improved resource use efficiency and stability of crop yields.

Plant metal chaperones: a novel perspective in dementia therapy.

Dys-homeostasis of copper metabolism and oxidative stress are major hallmarks in the brains of Alzheimer patients. Therefore, metal bioavailability and mechanisms of copper ion homeostasis throughout the body are crucial and potential targets for therapeutic agents. Many of the medications used or suggested, respectively, at present time, may either be toxic, reveal a lack of specificity or have unknown mechanisms of action in vivo. Metal chaperones from medicinal plants are proposed as medications that are relatively free from these disadvantages. Furthermore, these agents are a promising class of molecules for studies aimed at developing innovative and etiological treatments for protein-misfolding diseases, especially Alzheimer's disease.

Temperature responses of roots: impact on growth, root system architecture and implications for phenotyping.

Root phenotyping is a challenging task, mainly because of the hidden nature of this organ. Only recently, imaging technologies have become available that allow us to elucidate the dynamic establishment of root structure and function in the soil. In root tips, optical analysis of the relative elemental growth rates in root expansion zones of hydroponically-grown plants revealed that it is the maximum intensity of cellular growth processes rather than the length of the root growth zone that control the acclimation to dynamic changes in temperature. Acclimation of entire root systems was studied at high throughput in agar-filled Petri dishes. In the present study, optical analysis of root system architecture showed that low temperature induced smaller branching angles between primary and lateral roots, which caused a reduction in the volume that roots access at lower temperature. Simulation of temperature gradients similar to natural soil conditions led to differential responses in basal and apical parts of the root system, and significantly affected the entire root system. These results were supported by first data on the response of root structure and carbon transport to different root zone temperatures. These data were acquired by combined magnetic resonance imaging (MRI) and positron emission tomography (PET). They indicate acclimation of root structure and geometry to temperature and preferential accumulation of carbon near the root tip at low root zone temperatures. Overall, this study demonstrated the value of combining different phenotyping technologies that analyse processes at different spatial and temporal scales. Only such an integrated approach allows us to connect differences between genotypes obtained in artificial high throughput conditions with specific characteristics relevant for field performance. Thus, novel routes may be opened up for improved plant breeding as well as for mechanistic understanding of root structure and function.

Improved separation of palladium species in biological matrices by using a combination of gel permeation chromatography and isotachophoresis.

The binding of palladium to high-molecular-mass compounds in palladium-treated lettuce is investigated as an example for a biological matrix. The total palladium concentration in lettuce leaves is 10.3 ng/g wet weight. After homogenization, high-molecular-mass compounds (> 10 kDa) are isolated by ultrafiltration. For separation of these palladium species a combination of preparative gel permeation chromatography (GPC) and preparative isotachophoresis (ITP) is used. Palladium is determined in separated fractions by using a highly sensitive total reflection X-ray fluorescence (TXRF) method after preconcentration. After GPC separation, four main fractions of palladium species are collected, each containing palladium in ng quantities (3-10 ng). Two of these fractions are further separated by ITP, yielding at least three main peaks per GPC fraction, each containing palladium in the range of 0.3-3 ng. These palladium containing peaks are characterized by high-performance size exclusion chromatography (HPSEC) and capillary isotachophoresis (cITP) in parallel. HPSEC enables the estimation of the molecular mass of six main palladium peaks, covering a molecular mass range of 69-200 kDa. It is also shown that the estimation of molecular mass after separation is more reliable than the respective estimation directly in the first GPC run. However, cITP reveals that each of the separated peaks is still a mixture of at least five different compounds.