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Biomedical subjects

Anna Amtmann

Publications and source records attributed to Anna Amtmann.

10 recordsLinked to original sources

Low unidirectional sodium influx into root cells restricts net sodium accumulation in Thellungiella halophila, a salt-tolerant relative of Arabidopsis thaliana.

Thellungiella halophila is a useful model species for research into plant salt tolerance. It is closely related to Arabidopsis thaliana, but shows considerably higher salt tolerance. Comparative analysis of ion homeostasis in the two species allows the identification of ion transport pathways that are critical for salt tolerance and provides the basis for future studies into their molecular features. Previous studies indicated that salt tolerance in T. halophila is accompanied by low accumulation of Na in the leaves. Kinetic analysis of net ion uptake over three days confirmed lower Na uptake and K loss in T. halophila compared with A. thaliana. Differential net Na uptake rates were still apparent after 6 weeks of salt treatment. To assess the contribution of unidirectional Na fluxes to net Na uptake, kinetic studies of (22)Na fluxes were carried out in both species. The results show that unidirectional root Na influx is significantly lower in salt-grown T. halophila plants than in A. thaliana exposed to the same level of salinity (100 mM). Quantitative comparison of unidirectional influx and net Na accumulation suggests that both species operate efficient Na efflux, which partly compensates for Na influx. Kinetic analysis of (22)Na efflux indicated higher root Na efflux in A. thaliana than in T. halophila. Thus A. thaliana appears to spend more energy on Na export while nevertheless accumulating more Na than T. halophila. It is proposed that limitation of Na influx is the main mechanism by which T. halophila secures low net Na accumulation in saline conditions. This strategy provides the basis for a positive balance between growth and net Na uptake rates, which is essential for survival in high salt.

Arabidopsis↗

Arabidopsis thaliana cyclic nucleotide gated channel 3 forms a non-selective ion transporter involved in germination and cation transport.

The Arabidopsis thaliana genome contains 20 cyclic nucleotide gated channel (CNGC) genes encoding putative non-selective ion channels. Classical and reverse genetic approaches have revealed that two members of this family (CNGC2 and CNGC4) play a role in plant defence responses whereas CNGC1 and CNGC10 may participate in heavy metal and cation transport. Yet, it remains to be resolved how the ion transport attributes of CNGCs are integrated into their physiological function. In this study, CNGC3 is characterized through heterologous expression, GUS- and GFP-reporter gene fusions, and by adopting a reverse genetics approach. A CNGC3-GFP fusion protein shows that it is mainly targeted to the plasma membrane. Promoter GUS studies demonstrate CNGC3 expression predominantly in the cortical and epidermal root cells, but also a ubiquitous presence in shoot tissues. Expression of CNGC3 in yeast indicates it can function as a Na(+) uptake and a K(+) uptake mechanism. cngc3 null mutations decreased seed germination in the presence of NaCl but not KCl. Relative to the wild type, mutant seedling growth is more resistant to the presence of toxic concentrations of NaCl and KCl. The ionic composition and ion uptake characteristics of wild-type and mutant seedlings suggests that the growth advantage in these conditions may be due to restricted ion influx in mutant plants, and that CNGC3 functions in the non-selective uptake of monovalent cations in Arabidopsis root tissue.

Arabidopsis↗

Pollen development and fertilization in Arabidopsis is dependent on the MALE GAMETOGENESIS IMPAIRED ANTHERS gene encoding a type V P-type ATPase.

In flowering plants, development of the haploid male gametophytes (pollen grains) takes place in a specialized structure called the anther. Successful pollen development, and thus reproduction, requires high secretory activity in both anther tissues and pollen. In this paper, we describe a novel member of the eukaryotic type V subfamily (P(5)) of P-type ATPase cation pumps, the MALE GAMETOGENESIS IMPAIRED ANTHERS (MIA) gene. MIA protein is highly abundant in the endoplasmic reticulum and small vesicles of developing pollen grains and tapetum cells. T-DNA insertional mutants of MIA suffer from imbalances in cation homeostasis and exhibit a severe reduction in fertility. Mutant microspores fail to separate from tetrads and pollen grains are fragile with an abnormal morphology and altered cell wall structure. Disruption of MIA affects expression of genes essential for secretion as well as a high number of genes encoding cell wall proteins and membrane transporters. MIA functionally complements a mutant in the P(5) ATPase homolog SPF1 from Saccharomyces cerevisiae, suggesting a common function for P(5) ATPases in single and multicellular organisms. Our results suggest that MIA is required in the secretory pathway for proper secretion of vesicle cargo to the plasma membrane.

ATP-Binding Cassette Transporters↗

Vector analysis as a fast and easy method to compare gene expression responses between different experimental backgrounds.

BACKGROUND: Gene expression studies increasingly compare expression responses between different experimental backgrounds (genetic, physiological, or phylogenetic). By focusing on dynamic responses rather than a direct comparison of static expression levels, this type of study allows a finer dissection of primary and secondary regulatory effects in the various backgrounds. Usually, results of such experiments are presented in the form of Venn diagrams, which are intuitive and visually appealing, but lack a statistical foundation. RESULTS: Here we introduce Vector Analysis (VA) as a simple, yet principled, approach to comparing expression responses in different experimental backgrounds. VA enables the automatic assignment of genes to response prototypes and provides statistical significance estimates to eliminate spurious response patterns. The application of VA to a real dataset, comparing nutrient starvation responses in wild type and mutant Arabidopsis plants, reveals that consistent patterns of expression behavior are present in the data and are reliably detected by the algorithm. CONCLUSION: Vector analysis is a flexible, easy-to-use technique to compare gene expression patterns in different experimental backgrounds. It compares favorably with the classical Venn diagram approach and can be implemented manually using spreadsheets, such as Excel, or automatically by using the supplied software.

Algorithms↗

The potassium-dependent transcriptome of Arabidopsis reveals a prominent role of jasmonic acid in nutrient signaling.

Full genome microarrays were used to assess transcriptional responses of Arabidopsis seedlings to changing external supply of the essential macronutrient potassium (K(+)). Rank product statistics and iterative group analysis were employed to identify differentially regulated genes and statistically significant coregulated sets of functionally related genes. The most prominent response was found for genes linked to the phytohormone jasmonic acid (JA). Transcript levels for the JA biosynthetic enzymes lipoxygenase, allene oxide synthase, and allene oxide cyclase were strongly increased during K(+) starvation and quickly decreased after K(+) resupply. A large number of well-known JA responsive genes showed the same expression profile, including genes involved in storage of amino acids (VSP), glucosinolate production (CYP79), polyamine biosynthesis (ADC2), and defense (PDF1.2). Our findings highlight a novel role of JA in nutrient signaling and stress management through a variety of physiological processes such as nutrient storage, recycling, and reallocation. Other highly significant K(+)-responsive genes discovered in our study encoded cell wall proteins (e.g. extensins and arabinogalactans) and ion transporters (e.g. the high-affinity K(+) transporter HAK5 and the nitrate transporter NRT2.1) as well as proteins with a putative role in Ca(2+) signaling (e.g. calmodulins). On the basis of our results, we propose candidate genes involved in K(+) perception and signaling as well as a network of molecular processes underlying plant adaptation to K(+) deficiency.

Arabidopsis↗

Rank products: a simple, yet powerful, new method to detect differentially regulated genes in replicated microarray experiments.

One of the main objectives in the analysis of microarray experiments is the identification of genes that are differentially expressed under two experimental conditions. This task is complicated by the noisiness of the data and the large number of genes that are examined simultaneously. Here, we present a novel technique for identifying differentially expressed genes that does not originate from a sophisticated statistical model but rather from an analysis of biological reasoning. The new technique, which is based on calculating rank products (RP) from replicate experiments, is fast and simple. At the same time, it provides a straightforward and statistically stringent way to determine the significance level for each gene and allows for the flexible control of the false-detection rate and familywise error rate in the multiple testing situation of a microarray experiment. We use the RP technique on three biological data sets and show that in each case it performs more reliably and consistently than the non-parametric t-test variant implemented in Tusher et al.'s significance analysis of microarrays (SAM). We also show that the RP results are reliable in highly noisy data. An analysis of the physiological function of the identified genes indicates that the RP approach is powerful for identifying biologically relevant expression changes. In addition, using RP can lead to a sharp reduction in the number of replicate experiments needed to obtain reproducible results.

Acute Disease↗

Graph-based iterative Group Analysis enhances microarray interpretation.

BACKGROUND: One of the most time-consuming tasks after performing a gene expression experiment is the biological interpretation of the results by identifying physiologically important associations between the differentially expressed genes. A large part of the relevant functional evidence can be represented in the form of graphs, e.g. metabolic and signaling pathways, protein interaction maps, shared GeneOntology annotations, or literature co-citation relations. Such graphs are easily constructed from available genome annotation data. The problem of biological interpretation can then be described as identifying the subgraphs showing the most significant patterns of gene expression. We applied a graph-based extension of our iterative Group Analysis (iGA) approach to obtain a statistically rigorous identification of the subgraphs of interest in any evidence graph. RESULTS: We validated the Graph-based iterative Group Analysis (GiGA) by applying it to the classic yeast diauxic shift experiment of DeRisi et al., using GeneOntology and metabolic network information. GiGA reliably identified and summarized all the biological processes discussed in the original publication. Visualization of the detected subgraphs allowed the convenient exploration of the results. The method also identified several processes that were not presented in the original paper but are of obvious relevance to the yeast starvation response. CONCLUSIONS: GiGA provides a fast and flexible delimitation of the most interesting areas in a microarray experiment, and leads to a considerable speed-up and improvement of the interpretation process.

Algorithms↗

Iterative Group Analysis (iGA): a simple tool to enhance sensitivity and facilitate interpretation of microarray experiments.

BACKGROUND: The biological interpretation of even a simple microarray experiment can be a challenging and highly complex task. Here we present a new method (Iterative Group Analysis) to facilitate, improve, and accelerate this process. RESULTS: Our Iterative Group Analysis approach (iGA) uses elementary statistics to identify those functional classes of genes that are significantly changed in an experiment and at the same time determines which of the class members are most likely to be differentially expressed. iGA does not require that all members of a class change and is therefore robust against imperfect class assignments, which can be derived from public sources (e.g. GeneOntologies) or automated processes (e.g. key word extraction from gene names). In contrast to previous non-iterative approaches, iGA does not depend on the availability of fixed lists of differentially expressed genes, and thus can be used to increase the sensitivity of gene detection especially in very noisy or small data sets. In the extreme, iGA can even produce statistically meaningful results without any experimental replication. The automated functional annotation provided by iGA greatly reduces the complexity of microarray results and facilitates the interpretation process. In addition, iGA can be used as a fast and efficient tool for the platform-independent comparison of a microarray experiment to the vast number of published results, automatically highlighting shared genes of potential interest. CONCLUSIONS: By applying iGA to a wide variety of data from diverse organisms and platforms we show that this approach enhances and accelerates the interpretation of microarray experiments.

Animals↗

Transcriptome analysis of root transporters reveals participation of multiple gene families in the response to cation stress.

Plant nutrition critically depends on the activity of membrane transporters that translocate minerals from the soil into the plant and are responsible for their intra- and intercellular distribution. Most plant membrane transporters are encoded by multigene families whose members often exhibit overlapping expression patterns and a high degree of sequence homology. Furthermore, many inorganic nutrients are transported by more than one transporter family. These considerations, coupled with a large number of so-far non-annotated putative transporter genes, hamper our progress in understanding how the activity of specific transporters is integrated into a response to fluctuating conditions. We designed an oligonucleotide microarray representing 1096 Arabidopsis transporter genes and analysed the root transporter transcriptome over a 96-h period with respect to 80 mM NaCl, K+ starvation and Ca2+ starvation. Our data show that cation stress led to changes in transcript level of many genes across most transporter gene families. Analysis of transcriptionally modulated genes across all functional groups of transporters revealed families such as V-type ATPases and aquaporins that responded to all treatments, and families - which included putative non-selective cation channels for the NaCl treatment and metal transporters for Ca2+ starvation conditions - that responded to specific ionic environments. Several gene families including primary pumps, antiporters and aquaporins were analysed in detail with respect to the mRNA levels of different isoforms during ion stress. Cluster analysis allowed identification of distinct expression profiles, and several novel putative regulatory motifs were discovered within sets of co-expressed genes.

Arabidopsis↗