GWAS paper on Cd accumulation

The Salt lab recently published a paper in PLoS Genetics in which we describe the use of GWA mapping to identify HMA3 as the primary locus controlling natural variation in foliar accumulation of Cd in Arabidopsis thaliana. The paper can be read at http://www.plosgenetics.org/article/info%3Adoi%2F10.1371%2Fjournal.pgen.1002923 and the abstract can be seen below.

Abstract: Understanding the mechanism of cadmium (Cd) accumulation in plants is important to help reduce its potential toxicity to both plants and humans through dietary and environmental exposure. Here, we report on a study to uncover the genetic basis underlying natural variation in Cd accumulation in a world-wide collection of 349 wild collected Arabidopsis thaliana accessions. We identified a 4-fold variation (0.5-2 µg Cd g(-1) dry weight) in leaf Cd accumulation when these accessions were grown in a controlled common garden. By combining genome-wide association mapping, linkage mapping in an experimental F2 population, and transgenic complementation, we reveal that HMA3 is the sole major locus responsible for the variation in leaf Cd accumulation we observe in this diverse population of A. thaliana accessions. Analysis of the predicted amino acid sequence of HMA3 from 149 A. thaliana accessions reveals the existence of 10 major natural protein haplotypes. Association of these haplotypes with leaf Cd accumulation and genetics complementation experiments indicate that 5 of these haplotypes are active and 5 are inactive, and that elevated leaf Cd accumulation is associated with the reduced function of HMA3 caused by a nonsense mutation and polymorphisms that change two specific amino acids.

New Perspective in Science

Brian Dilkes and I have a perspective in the new issue of Science titled "Elemental Profiles Reflect Plant Adaptations to the Environment". The abstract is below. Science puts a very (and I mean VERY) tight word limit on the manuscript and so one of the things that we needed to cut was the acknowledgements. So we would like to take the time here to give huge thanks to all the great people who took the time to read drafts and give us (lots of) feedback. 

Jim Fleet,David Salt, Kirsten Bomblies, Elizabeth Haswell, Elizabeth Buescher, Nancy Emery, Clint Chapple, Jody Banks, Joanna Dinsmore, Luca Comai, Aimee Terauchi, Greg Ziegler and five anonymous reviewers for comments on the perspective. 

Pamela Hines, our editor at Science, contributed lots of wisdom, insight and most of all patience, for which we are ever grateful. 

THANKS!

Ivan

Most mineral elements found in plant tissues come exclusively from the soil, necessitating that plants adapt to highly variable soil compositions to survive and thrive. Profiling element concentrations in genetically diverse plant populations is providing insights into the plant-environment interactions that control elemental accumulation, as well as identifying the underlying genes. The resulting molecular understanding of plant adaptation to the environment both demonstrates how soils can shape genetic diversity and provides solutions to important agricultural challenges.                      

 

New Paper in PLoS ONE

Our most recent paper was just published in PLoS ONE. "Biodiversity of Mineral Nutrient and Trace Element Accumulation in Arabidopsis thaliana" is our first installment of all the association panel data that we have generated.  It looks at the corrections between elements within and between tissues in the initial 96 accession 'Nordborg' panel.  It includes some very nice data on hydroponics grown plants from samples generated by Christian Hermans and Nathalie Verbruggen from Université Libre de Bruxelles in Belgium.  Special thanks are due to Jennie Hard for all the help with figure preparation. There will be several more papers analyzing this and similar data submitted in the (hopefully) near future.  

 

Here is the abstract:

In order to grow on soils that vary widely in chemical composition, plants have evolved mechanisms for regulating the elemental composition of their tissues to balance the mineral nutrient and trace element bioavailability in the soil with the requirements of the plant for growth and development. The biodiversity that exists within a species can be utilized to investigate how regulatory mechanisms of individual elements interact and to identify genes important for these processes. We analyzed the elemental composition (ionome) of a set of 96 wild accessions of the genetic model plant Arabidopsis thaliana grown in hydroponic culture and soil using inductively coupled plasma mass spectrometry (ICP-MS). The concentrations of 17–19 elements were analyzed in roots and leaves from plants grown hydroponically, and leaves and seeds from plants grown in artificial soil. Significant genetic effects were detected for almost every element analyzed. We observed very few correlations between the elemental composition of the leaves and either the roots or seeds. There were many pairs of elements that were significantly correlated with each other within a tissue, but almost none of these pairs were consistently correlated across tissues and growth conditions, a phenomenon observed in several previous studies. These results suggest that the ionome of a plant tissue is variable, yet tightly controlled by genes and gene×environment interactions. The dataset provides a valuable resource for mapping studies to identify genes regulating elemental accumulation. All of the ionomic data is available at www.ionomicshub.org.

 

and here is the non-technical summary:

Understanding how plants regulate element composition of tissues is critical for agriculture, the environment, and human health. Sustainably meeting the increasing food and biofuel demands of the planet will require growing crops with fewer inputs such as the primary macronutrients phosphorus (P) and potassium (K). Ionomics is the study of elemental accumulation in living systems using high-throughput elemental profiling. With this technique, we can rapidly generate large quantities of data on thousands of samples, allowing for the profiling of large genetic mapping populations and the discovery of hundreds of loci important for elemental accumulation. We have used this approach to sample the natural diversity present in collections of a model plant, the wild mustard Arabidopsis. We find that the elemental composition of a plant is tightly controlled by its genes. We also find that elements will have different relationships between them depending on the environment and the tissues (root, seed or leaf) under study. This suggests that crop varieties developed for improved elemental uptake and accumulation will be highly environment specific.

First iHUB workshop

The first NSF sponsored iHUB workshop held 26 & 27 April, 2012 in St Louis, US was a success. New tools and features discussed at the workshop should be appearing on the iHUB soon.

Aberdeen Ionomics Facility running

The ICP-MS instruments in David Salt's Ionomics Facility at the University of Aberdeen are now up and running.

PLoS one paper on CPR5 and potassium

The Salt group has published a new paper on a novel role of CPR5 in the regulation of potassium in A. thaliana.

Borghi M, Rus A, Salt DE (2011) Loss-of-Function of Constitutive Expresser of Pathogenesis Related Genes5 Affects Potassium Homeostasis in Arabidopsis thaliana. PLoS ONE 6(10): e26360. doi:10.1371/journal.pone.0026360

Here is the Abstract
Here, we demonstrate that the reduction in leaf K⁺ observed in a mutant previously identified in an ionomic screen of fast neutron mutagenized Arabidopsis thaliana is caused by a loss-of-function allele of CPR5, which we name cpr5-3. This observation establishes low leaf K⁺ as a new phenotype for loss-of-function alleles of CPR5. We investigate the factors affecting this low leaf K⁺ in cpr5 using double mutants defective in salicylic acid (SA) and jasmonic acid (JA) signalling, and by gene expression analysis of various channels and transporters. Reciprocal grafting between cpr5 and Col-0 was used to determine the relative importance of the shoot and root in causing the low leaf K⁺ phenotype of cpr5. Our data show that loss-of-function of CPR5 in shoots primarily determines the low leaf K⁺ phenotype of cpr5, though the roots also contribute to a lesser degree. The low leaf K⁺ phenotype of cpr5 is independent of the elevated SA and JA known to occur in cpr5. In cpr5 expression of genes encoding various Cyclic Nucleotide Gated Channels (CNGCs) are uniquely elevated in leaves. Further, expression of HAK5, encoding the high affinity K⁺ uptake transporter, is reduced in roots of cpr5 grown with high or low K⁺ supply. We suggest a model in which low leaf K⁺ in cpr5 is driven primarily by enhanced shoot-to-root K⁺ export caused by a constitutive activation of the expression of various CNGCs. This activation may enhance K⁺ efflux, either indirectly via enhanced cytosolic Ca²⁺ and/or directly by increased K⁺ transport activity. Enhanced shoot-to-root K⁺ export may also cause the reduced expression of HAK5 observed in roots of cpr5, leading to a reduction in uptake of K⁺. All ionomic data presented is publically available at www.ionomicshub.org.

Two new Plant Cell Papers

Two new papers have been published in Plant Cell in the last month describing genes that we cloned from the original Lahner et al. ionomics screen:

 

The first is "Arabidopsis NPCC6/NaKR1 Is a Phloem Mobile Metal Binding Protein Necessary for Phloem Function and Root Meristem Maintenance".  This paper was mainly the result of the hard work of newly minted Ph.D Hui Tian from John Ward's Lab at UMN.  She worked with us to clone the gene, finally finding it when the causal deletion of only 7 bp disrupted a single oligo on the Arabidopsis tiling array. It's a fascinating gene, encoding a protein that moves through the phloem, the part of the plants vasculature responsible for moving solutes away from leaves.

The second is "Sphingolipids in the Root Play an Important Role in Regulating the Leaf Ionome in Arabidopsis thaliana".  A great collaboration between our group and several groups working on sphingolipids resulted when we landed on a gene in the sphingolipid pathway. Subtly altering the sphingolipid pathway results in what appears to be two different ionomics associated phenotypes: altered suberin and Fe homeostasis.

 

Here is the abstract of the first paper:

SODIUM POTASSIUM ROOT DEFECTIVE1 (NaKR1; previously called NPCC6)encodes a soluble metal binding protein that is specificallyexpressed in companion cells of the phloem. The nakr1-1 mutantphenotype includes high Na⁺, K⁺, Rb⁺, and starch accumulationin leaves, short roots, late flowering, and decreased long-distancetransport of sucrose. Using traditional and DNA microarray-baseddeletion mapping, a 7-bp deletion was found in an exon of NaKR1that introduced a premature stop codon. The mutant phenotypeswere complemented by transformation with the native gene orNaKR1-GFP (green fluorescent protein) and NaKR1-β-glucuronidasefusions driven by the native promoter. NAKR1-GFP was mobilein the phloem; it moved from companion cells into sieve elementsand into a previously undiscovered symplasmic domain in theroot meristem. Grafting experiments revealed that the high Na⁺accumulation was due mainly to loss of NaKR1 function in theleaves. This supports a role for the phloem in recirculatingNa⁺ to the roots to limit Na⁺ accumulation in leaves. The onsetof root phenotypes coincided with NaKR1 expression after germination.The nakr1-1 short root phenotype was due primarily to a decreasedcell division rate in the root meristem, indicating a role inroot meristem maintenance for NaKR1 expression in the phloem.

And here is a summary intended for lay audiences:

A major problem for world agriculture is the growing decrease in avaialable arable land. More and more we are working in solils that impart a stress on the plants that make up the crops we depend on. In order for plants to survive without being able to move out of unfavorable soil environments, they adjust the biochemical composition of their tissues through a wide variety of mechanisms.  One of these mechanisms is to move  elements such as sodium (Na) and potassium (K) from tissue to tissue, including from the root to the shoot and back again.  Understanding the molecular basis of these mechanisms will enable the production of crops that are better able to respond to the changing environment  and increase yields with fewer inputs.  In this study, we identified and characterized a gene which is important for loading Na  into the phloem, the 'veins' of the plant responsible for moving molecules out of the leaves to the seeds and roots. The protein also moves into the  phloem. Plants without a functional form of this gene, called NAKR1, have altered levels of Na, K and starch in the leaves, have shorter roots and flower later than plants with a functional copy of NAKR1.  These results will lead to a better understanding of how plants distribute elements between tissues and ultimately will allow for crop improvement strategies that deal with poor soil quality.

 

And here is the abstract of the second paper:

Sphingolipid synthesis is initiated by condensation of Ser with palmitoyl-CoA producing 3-ketodihydrosphinganine (3-KDS), which is reduced by a 3-KDS reductase to dihydrosphinganine. Ser palmitoyltransferase is essential for plant viability. Arabidopsis thaliana contains two genes (At3g06060/TSC10A and At5g19200/TSC10B) encoding proteins with significant similarity to the yeast 3-KDS reductase, Tsc10p. Heterologous expression in yeast of either Arabidopsis gene restored 3-KDS reductase activity to the yeast tsc10Δ mutant, confirming both as bona fide 3-KDS reductase genes. Consistent with sphingolipids having essential functions in plants, double mutant progeny lacking both genes were not recovered from crosses of single tsc10A and tsc10B mutants. Although the 3-KDS reductase genes are functionally redundant and ubiquitously expressed in Arabidopsis, 3-KDS reductase activity was reduced to 10% of wild-type levels in the loss-of-function tsc10a mutant, leading to an altered sphingolipid profile. This perturbation of sphingolipid biosynthesis in the Arabidopsis tsc10a mutant leads an altered leaf ionome, including increases in Na, K, and Rb and decreases in Mg, Ca, Fe, and Mo. Reciprocal grafting revealed that these changes in the leaf ionome are driven by the root and are associated with increases in root suberin and alterations in Fe homeostasis.

And here is a summary intended for lay audiences:

Sphingolipids, a class of membrane lipids with essential functions in all Eukaryotes, are thought to make up a large percentage of some plant membranes and have specific roles in cell processes through the formation of small microdomains. Here we discuss the role of two genes in the sphigolipid biosynthesis pathway in the model plant Arabidopsis Thaliana.  When both genes are disrupted, the plants are not viable. However, when the higer expressed gene  is disrupted, the  plants look normal but elemental profiling reveals that they have significantly altered elemental accumulation in their leaves.  Several of the changes appear to be the result of altering the the amount of suberin, a polymer which forms a barrier to water and ion movement in the root, is altered.  We also observed alterations in the plants Fe homestasis mechanisms, the cause of which  is still unknown. Understanding these processes will enable the prodcution of crops that are more efficient in their water and nutrient uptake effficiency.