The human genome holds an extraordinary trove of information about human development, physiology, medicine and evolution. Here we report the results of an international collaboration to produce and make freely available a draft sequence of the human genome. We also present an initial analysis of the data, describing some of the insights that can be gleaned from the sequence.

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Initial sequencing and analysis of the human genome.

Salt and drought stress signal transduction consists of ionic and osmotic homeostasis signaling pathways, detoxification (i.e., damage control and repair) response pathways, and pathways for growth regulation. The ionic aspect of salt stress is signaled via the SOS pathway where a calcium-responsive SOS3-SOS2 protein kinase complex controls the expression and activity of ion transporters such as SOS1. Osmotic stress activates several protein kinases including mitogen-activated kinases, which may mediate osmotic homeostasis and/or detoxification responses. A number of phospholipid systems are activated by osmotic stress, generating a diverse array of messenger molecules, some of which may function upstream of the osmotic stress-activated protein kinases. Abscisic acid biosynthesis is regulated by osmotic stress at multiple steps. Both ABA-dependent and -independent osmotic stress signaling first modify constitutively expressed transcription factors, leading to the expression of early response transcriptional activators, which then activate downstream stress tolerance effector genes.

/pdf/salt-and-drought-stress-signal-transduction-in-plants-4bj87ki2ww.pdf

Salt and drought stress signal transduction in plants

We explored genomic expression patterns in the yeast Saccharomyces cerevisiae responding to diverse environmental transitions. DNA microarrays were used to measure changes in transcript levels over time for almost every yeast gene, as cells responded to temperature shocks, hydrogen peroxide, the superoxide-generating drug menadione, the sulfhydryl-oxidizing agent diamide, the disulfide-reducing agent dithiothreitol, hyper- and hypo-osmotic shock, amino acid starvation, nitrogen source depletion, and progression into stationary phase. A large set of genes (approximately 900) showed a similar drastic response to almost all of these environmental changes. Additional features of the genomic responses were specialized for specific conditions. Promoter analysis and subsequent characterization of the responses of mutant strains implicated the transcription factors Yap1p, as well as Msn2p and Msn4p, in mediating specific features of the transcriptional response, while the identification of novel sequence elements provided clues to novel regulators. Physiological themes in the genomic responses to specific environmental stresses provided insights into the effects of those stresses on the cell.

/pdf/genomic-expression-programs-in-the-response-of-yeast-cells-4xrnicxf7t.pdf

Genomic expression programs in the response of yeast cells to environmental changes.

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▪ Abstract Plant responses to salinity stress are reviewed with emphasis on molecular mechanisms of signal transduction and on the physiological consequences of altered gene expression that affect biochemical reactions downstream of stress sensing. We make extensive use of comparisons with model organisms, halophytic plants, and yeast, which provide a paradigm for many responses to salinity exhibited by stress-sensitive plants. Among biochemical responses, we emphasize osmolyte biosynthesis and function, water flux control, and membrane transport of ions for maintenance and re-establishment of homeostasis. The advances in understanding the effectiveness of stress responses, and distinctions between pathology and adaptive advantage, are increasingly based on transgenic plant and mutant analyses, in particular the analysis of Arabidopsis mutants defective in elements of stress signal transduction pathways. We summarize evidence for plant stress signaling systems, some of which have components analogous to t...

Plant cellular and molecular responses to high salinity.

MAP kinase pathways are conserved signalling systems in eukaryotes that control stress responses, cell growth, and proliferation, as well as differentiation. Here, we discuss and compare the feedback control mechanisms of two very well studied yeast signalling systems: the pheromone response pathway and the osmosensing HOG pathway. Mathematical models have recently been generated, allowing in silico analysis of signalling properties of both pathways. To advance our understanding of pathway control and to make modelling less dependent on parameter estimation, quantitative time course data of high precision and resolution need to be generated in the future and implemented into mathematical models. We expect that a combination of quantitative analyses and modelling/simulation will provide novel insight into the rules with which signalling pathways control cellular processes.

Modelling signalling pathways-A yeast approach

We investigated the importance of the interaction between the N1‘-atom of the cofactor thiamine diphosphate and glutamic acid residue 51 in pyruvate decarboxylase (EC 4.1.1.1). The yeast wild type gene PDC1 and the respective mutant genes (E51Q and E51A) were expressed in Escherichia coli. The three enzymes were purified to homogeneity. They comigrated as a single band during silver-stained SDS/PAGE with a molecular mass of 60 000 Da. A molecular mass of 61 200 ± 200 Da was determined by mass spectrometry for the subunit. The native enzyme is a homotetramer as demonstrated by gel filtration experiments. Near- and far-UV CD spectra showed no significant differences for the apoenzyme of the wild type and the mutants. Slight differences in the rate of thiamine diphosphate binding to the apoprotein component were observed between the wild type and the E51Q PDC by CD spectroscopy. Compared to the wild type enzyme, thiamine diphosphate binding at the E51A mutant apoprotein is very slow. Only 0.04% of the cataly...

Role of Glu51 for cofactor binding and catalytic activity in pyruvate decarboxylase from yeast studied by site-directed mutagenesis.

We present a simple ordinary differential equation (ODE) model of the adaptive response to an osmotic shock in the yeast Saccharomyces cerevisiae. The model consists of two main components. First, a biophysical model describing how the cell volume and the turgor pressure are affected by varying extra-cellular osmolarity. The second component describes how the cell controls the biophysical system in order to keep turgor pressure, or equivalently volume, constant. This is done by adjusting the glycerol production and the glycerol outflow from the cell. The complete model consists of 4 ODEs, 3 algebraic equations and 10 parameters. The parameters are constrained from various literature sources and estimated from new and previously published absolute time series data on intra-cellular and total glycerol. The qualitative behaviour of the model has been successfully tested on data from other genetically modified strains as well as data for different input signals. Compared to a previous detailed model of osmoregulation, the main strength of our model is its lower complexity, contributing to a better understanding of osmoregulation by focusing on relationships which are obscured in the more detailed model. Besides, the low complexity makes it possible to obtain more reliable parameter estimates.

A simple mathematical model of adaptation to high osmolarity in yeast

The Saccharomyces cerevisiae strain Σ1278b possesses two putative aquaporins, Aqy1-1p and Aqy2-1p. Previous work demonstrated that Aqy1-1p functions as a water channel in Xenopus oocyte. However, no function could be attributed to Aqy2-1p in this system. Specific antibodies were used to follow the expression of Aqy1-1p and Aqy2-1p in the yeast. Aqy1-1p was never detected whatever the growth phase and culture conditions tested. In contrast, Aqy2-1p was detected only during the exponential growth phase in rich medium containing glucose. Aqy2-1p expression was repressed by hyper-osmotic culture conditions. Both immunocytochemistry and biochemical subcellular fractionation demonstrated that Aqy2-1p is located on the endoplasmic reticulum (ER) as well as on the plasma membrane. In microsomal vesicles enriched in ER, a water channel activity due to Aqy2-1p was detected by stopped-flow analysis. Our results show that the expression of aquaporins is tightly controlled. The physiological relevance of aquaporin-mediated water transport in yeast is discussed.

Existence of a tightly regulated water channel in Saccharomyces cerevisiae.

Comparative genomics allows comparison of different proteins that execute presumably identical functions in different organisms. In contrast to paralogues, orthologues per definition perform the same function and interact with the same partners and, consequently, should display conservation in all these properties. We have employed 20 fungal genomes to analyse key components of the high osmolarity glycerol signalling pathway of Saccharomyces cerevisiae. Among the proteins scrutinised are a complete phosphotransfer module, a MAP kinase, two scaffold proteins, one of which is also a MAPKK, and two transcription factors. Sequence alignments, domain structure and size analysis, combined with the rich information available in the literature, allowed us to probe previous structural and functional studies and to generate hypotheses for future experimental studies. Although certain domains are too highly conserved across fungal species for meaningful comparative studies, others, like interaction domains, can be studied in closely related species. Moreover, putative functionally relevant sites for protein modifications can be identified in such comparative studies. We provide several relevant examples and present a number of previously un(der)characterised domains of potential functional significance in osmosensing and signal transduction. We propose that any functional protein analysis in fungi should make use of the unique resource that fungal genome sequences offer.

Stefan Hohmann

Papers

Modelling signalling pathways-A yeast approach

Role of Glu51 for cofactor binding and catalytic activity in pyruvate decarboxylase from yeast studied by site-directed mutagenesis.

A simple mathematical model of adaptation to high osmolarity in yeast

Existence of a tightly regulated water channel in Saccharomyces cerevisiae.

Comparative analysis of HOG pathway proteins to generate hypotheses for functional analysis