Environmental genetics of root system architecture

The root system is the plant’s principal organ for water and mineral nutrient supply. Root growth follows an endogenous, developmental programme. Yet, this programme can be modulated by external cues which makes root system architecture (RSA), the spatial configuration of all root parts, a highly plastic trait. Presence or absence of nutrients such as nitrate (N), phosphate (P), potassium (K) and sulphate (S) serve as environmental signals to which a plant responds with targeted proliferation or restriction of main or lateral root growth. In turn, RSA serves as a quantitative reporter system of nutrient starvation responses and can therefore be used to study nutrient sensing and signalling mechanisms. In this study, I have analysed root architectural responses of various Arabidopsis thaliana genotypes (wildtype, mutants and natural accessions) to single and multiple nutrient deficiency treatments. A comprehensive analysis of combinatorial N, P, K an S supply allowed me to dissect the effect of individual nutrients on individual root parameters. It also highlighted the existence of interactive effects arising from simultaneous environmental stimuli. Quantification of appropriate RSA parameters allowed for targeted testing of known regulatory genes in specific nutritional settings. This revealed, for example, a novel role for CIPK23, AKT1 and NRT1.1 in integrating K and N effects on higher order lateral root branching and main root angle. A significant contribution to phenotypic variation also arose from P*K interactions. I could show that the iron (Fe) concentration in the external medium is an important driving force of RSA responses to low-P and low-K. In fact, P and K deprivation caused Fe accumulation in distinct parts of the root system, as demonstrated by Fe staining and synchrotron X-Ray fluorescence. Again, selected K, P and Fe transport and signalling mutants were tested for aberrant low-K and/or low-P phenotypes. Most notably, the two paralogous ER-localised multicopper oxidases LPR1 and LPR2 emerged as important signalling components of P and K deprivation, potentially integrating Fe homeostasis with meristematic activity under these conditions. In addition to the targeted characterisation of specific genotype-environment interactions, I investigated novel RSA responses to low-K via a non-targeted approach based on natural variation. A morphological gradient spanned the entire genotype set, linking two extreme strategies of low-K responses. Strategy I accessions responded to low-K with a moderate reduction of main root growth but a severe restriction of lateral root elongation. In contrast, strategy II genotypes ceded main root growth in favour of lateral root proliferation. The genetic basis of these low-K responses was then subsequently mapped onto the A. thaliana genome via quantitative trait loci (QTL) analysis using recombinant inbred lines derived from parental accessions that either adopt strategy I (Col-0) or II (Ct-1). In sum, this study addresses the question how plants incorporate environmental signals to modulate developmental programmes that underly RSA formation. I present evidence for novel phenotypic responses to nutrient deprivation and for novel genetic regulators involved in nutrient signalling and crosstalk

Diffusion and Precipitation Processes in Iron-Based Silica Gardens

Silica gardens are tubular structures that form along the interface of multivalent metal salts and alk. solns. of sodium silicate, driven by a complex interplay of osmotic and buoyant forces together with chem. reaction. They display peculiar plant-​like morphologies and thus can be considered as one of the few examples for the spontaneous biomimetic self-​ordering of purely inorg. materials. Recently, we could show that silica gardens moreover are highly dynamic systems that remain far from equil. for considerable periods of time long after macroscopic growth is completed. Due to initial compartmentalization, drastic concn. gradients were found to exist across the tube walls, which give rise to noticeable electrochem. potential differences and decay only slowly in a series of coupled diffusion and pptn. processes. The effect of the nature of the metal cations on the dynamic behavior of the system has been studied. The authors have grown single macroscopic silica garden tubes by controlled addn. of sodium silicate sol to pellets of iron(II) and iron(III) chloride. In the following, the concns. of ionic species were measured as a function of time on both sides of the formed membranes, while electrochem. potentials and pH were monitored online by immersing the corresponding sensors into the two sepd. soln. reservoirs. At the end of the expts., the solid tube material was furthermore characterized with respect to compn. and microstructure by a combination of ex situ techniques. The collected data are compared to the previously reported case of cobalt-​based silica gardens and used to shed light on ion diffusion through the inorg. membranes as well as progressive mineralization at both surfaces of the tube walls. These results reveal important differences in the dynamics of the three studied systems, which can be explained based on the acidity of the metal cations and the porosity of the membranes, leading to substantially dissimilar time-​dependent soln. chem. as well as distinct final mineral structures. The insight gained in this work may help to better understand the diffusion properties and pptn. patterns in tubular iron (hydr)​oxide​/silicate structures obsd. in geol. environments and during steel corrosion

Isotope Ratio Outlier Analysis (IROA) for HPLC–TOFMS-Based Metabolomics of Human Urine

Metabolic fingerprinting by mass spectrometry aims at the comprehensive, semiquantitative analysis of metabolites. Isotope dilution, if successfully implemented, may provide a more reliable, relative quantification. Therefore, the 13C labeled yeast extract of the IROA TruQuant kit was added as an internal standard (IS) to human urine samples measured in full-scan mode on a high-performance liquid chromatography-time-of-flight mass spectrometer (HPLC–TOFMS) system. The isotope ratio approach enabled the analysis of 112 metabolites. The correlation with reference data did not improve significantly using 12C/13C ratios compared to absolute 12C peak areas. Moreover, using an intricate 13C-labeled standard increased the complexity of the mass spectra, which made correct signal annotation more challenging. On the positive side, the ratio approach helps to reduce batch effects, but it does not perform better than computational methods such as the “removebatcheffect” function in the R package Limma

MCT4 blockade increases the efficacy of immune checkpoint blockade

Background & Aims Intratumoral lactate accumulation and acidosis impair T-cell function and antitumor immunity. Interestingly, expression of the lactate transporter monocarboxylate transporter (MCT) 4, but not MCT1, turned out to be prognostic for the survival of patients with rectal cancer, indicating that single MCT4 blockade might be a promising strategy to overcome glycolysis-related therapy resistance. Methods To determine whether blockade of MCT4 alone is sufficient to improve the efficacy of immune checkpoint blockade (ICB) therapy, we examined the effects of the selective MCT1 inhibitor AZD3965 and a novel MCT4 inhibitor in a colorectal carcinoma (CRC) tumor spheroid model co-cultured with blood leukocytes in vitro and the MC38 murine CRC model in vivo in combination with an antibody against programmed cell death ligand-1(PD-L1). Results Inhibition of MCT4 was sufficient to reduce lactate efflux in three-dimensional (3D) CRC spheroids but not in two-dimensional cell-cultures. Co-administration of the MCT4 inhibitor and ICB augmented immune cell infiltration, T-cell function and decreased CRC spheroid viability in a 3D co-culture model of human CRC spheroids with blood leukocytes. Accordingly, combination of MCT4 and ICB increased intratumoral pH, improved leukocyte infiltration and T-cell activation, delayed tumor growth, and prolonged survival in vivo. MCT1 inhibition exerted no further beneficial impact. Conclusions These findings demonstrate that single MCT4 inhibition represents a novel therapeutic approach to reverse lactic-acid driven immunosuppression and might be suitable to improve ICB efficacy

Additive-induced morphological tuning of self-assembled silica-barium carbonate crystal aggregates

Crystallisation of barium carbonate from alkaline silica solutions results in the formation of extraordinary micron-scale architectures exhibiting non-crystallographic curved shapes, such as helical filaments and worm-like braids. These so-called "silic

EZ-Root-VIS: A Software Pipeline for the Rapid Analysis and Visual Reconstruction of Root System Architecture

If we want to understand how the environment has shaped the appearance and behavior of living creatures, we need to compare groups of individuals that differ in genetic makeup and environment experience. For complex phenotypic features, such as body posture or facial expression in humans, comparison is not straightforward because some of the contributing factors cannot easily be quantified or averaged across individuals. Therefore, computational methods are used to reconstruct representative prototypes using a range of algorithms for filling in missing information and calculating means. The same problem applies to the root system architecture (RSA) of plants. Several computer programs are available for extracting numerical data from root images, but they usually do not offer customized data analysis or visual reconstruction of RSA. We developed Root-VIS, a free software tool that facilitates the determination of means and variance of many different RSA features across user-selected sets of root images. Furthermore, Root-VIS offers several options to generate visual reconstructions of root systems from the averaged data to enable screening and modeling. We confirmed the suitability of Root-VIS, combined with a new version of EZ-Rhizo, for the rapid characterization of genotype-environment interactions and gene discovery through genome-wide association studies in Arabidopsis (Arabidopsis thaliana)

The effect of silica on polymorphic precipitation of calcium carbonate: an on-line energy-dispersive X-ray diffraction (EDXRD) study

Calcium carbonate is the most abundant biomineral and a compd. of great industrial importance. Its pptn. from soln. has been studied extensively and was often shown to proceed via distinct intermediate phases, which undergo sequential transformations before eventually yielding the stable cryst. polymorph, calcite. In the present work, we have investigated the crystn. of calcium carbonate in a time-resolved and non-invasive manner by means of energy-dispersive X-ray diffraction (EDXRD) using synchrotron radiation. In particular, the role of silica as a sol. additive during the crystn. process was examd. Measurements were carried out at different temps. (20, 50 and 80 °C) and various silica concns. Expts. conducted in the absence of silica reflect the continuous conversion of kinetically formed metastable polymorphs (vaterite and aragonite) to calcite and allow for quantifying the progress of transformation. Addn. of silica induced remarkable changes in the temporal evolution of polymorphic fractions existing in the system. Essentially, the formation of calcite was found to be accelerated at 20 °C, whereas marked retardation or complete inhibition of phase transitions was obsd. at higher temps. These findings are explained in terms of a competition between the promotional effect of silica on calcite growth rates and kinetic stabilization of vaterite and aragonite due to adsorption (or pptn.) of silica on their surfaces, along with temp.-dependent variations of silica condensation rates. Data collected at high silica concns. indicate the presence of an amorphous phase over extended frames of time, suggesting that initially generated ACC particles are progressively stabilized by silica. Our results may have important implications for CaCO3 pptn. scenarios in both geochem. and industrial settings, where soln. silicate is omnipresent, as well as for CO2 sequestration technologies. [on SciFinder(R)