Transcriptional profiles of primary metabolism and signal transduction-related genes in response to water stress in field-grown sunflower genotypes using a thematic cDNA microarray

A sunflower cDNA microarray containing about 800 clones covering major metabolic and signal transduction pathways was used to study gene expression profiles in leaves and embryos of drought-tolerant and -sensitive genotypes subjected to water-deficit stress under field conditions. Using two-step ANOVA normalization and analysis models, we identified 409 differentially expressed genes among genotypes, water treatment and organs. The majority of the cDNA clones differentially expressed under water stress was found to display opposite gene expression profiles in drought-tolerant genotype compared to drought-sensitive genotype. These dissimilarities suggest that the difference between tolerant and non-tolerant plants seems to be associated with changes in qualitative but not quantitative mRNA expression. Comparing leaves and embryos, 82 cDNA clones showing organ-specific variation in gene expression levels were identified in response to water stress across genotypes. Genes related to amino acids and carbohydrates metabolisms, and signal transduction were induced in embryos and repressed in leaves; suggesting that vegetative and reproductive organs respond differentially to water stress. Adaptive mechanisms controlling water deficit tolerance are proposed and discussed.     

酿酒酵母中SSK2基因与其他镉耐受相关基因之间的双基因缺失菌株构建

镉是一种严重的环境污染物,对人体具有致癌性,能蓄积在生物体内影响机体的生长、发育和生殖。有丝分裂原蛋白激酶(Mitogen-activated protein kinase,MAPK)在调节细胞存活、增殖和分化中是重要的信号分子,并能够被镉胁迫激活。酿酒酵母中2个MAPK信号传导途径,高渗透压甘油(High Osmolarity Glycerol,HOG)途径和细胞壁完整性(Cell Wall Integrity,CWI)途径都参与Cd2+胁迫下的细胞应答。为了进一步研究这两条途径在调控Cd2+胁迫方面的相互作用,以HOG途径的蛋白激酶SSK2基因为例,通过合成遗传阵列(Synthetic Genetic Array,SGA)方法,成功构建了SSK2基因与其他52个Cd2+耐受相关基因之间的双基因缺失菌株。为大规模研究Cd2+耐受基因之间在调控镉胁迫方面的遗传学相互作用奠定了基础,也为酿酒酵母的相关研究提供了一个新的遗传学手段。     

酵母HOG-MAPK途径

酿酒酵母Saccharomyces cerevisiae的高渗透性甘油促分裂原活化蛋白激酶(highos-molarity glycerol mitogen-activated protein kinase,HOG-MAPK)途径是高度保守的信号转导途径,很多方面和高等真核生物MAPK途径类似。该途径在高渗应激环境下控制信号转导和基因表达,是细胞生存所必需的。现对酵母HOG-MAPK途径的信号转导以及信号传递的专一性控制、HOG-MAPK途径各组分的亚细胞定位和基因表达调控机制进行综述。     

Reconstruction and in silico analysis of the MAPK signaling pathways in the human blood fluke, Schistosoma japonicum

At present, little is known about signal transduction mechanisms in schistosomes, which cause the disease of schistosomiasis. The mitogen-activated protein kinase (MAPK) signaling pathways, which are evolutionarily conserved from yeast to Homo sapiens, play key roles in multiple cellular processes. Here, we reconstructed the hypothetical MAPK signaling pathways in Schistosoma japonicum and compared the schistosome pathways with those of model eukaryote species. We identified 60 homologous components in the S. japonciumMAPK signaling pathways. Among these, 27 were predicted to be full-length sequences. Phylogenetic analysis of these proteins confirmed the evolutionary conservation of the MAPK signaling pathways. Remarkably, we identified S. japonicum homologues of GTP-binding protein beta and alpha-I subunits in the yeast mating pathway, which might be involved in the regulation of different life stages and female sexual maturation processes as well in schistosomes. In addition, several pathway member genes, including ERK, JNK, Sja-DSP, MRAS and RAS, were determined through quantitative PCR analysis to be expressed in a stage-specific manner, with ERK, JNK and their inhibitor Sja-DSP markedly upregulated in adult female schistosomes.     

Identification of mitogen-activated protein kinase signaling pathways that confer resistance to endoplasmic reticulum stress in Saccharomyces cerevisiae

Hypoxia activates all components of the unfolded protein response (UPR), a stress response initiated by the accumulation of unfolded proteins within the endoplasmic reticulum (ER). Our group and others have shown previously that the UPR, a hypoxia-inducible factor-independent signaling pathway, mediates cell survival during hypoxia and is required for tumor growth. Identifying new genes and pathways that are important for survival during ER stress may lead to the discovery of new targets in cancer therapy. Using the set of 4,728 homozygous diploid deletion mutants in budding yeast, Saccharomyces cerevisiae, we did a functional screen for genes that conferred resistance to ER stress-inducing agents. Deletion mutants in 56 genes showed increased sensitivity under ER stress conditions. Besides the classic UPR pathway and genes related to calcium homeostasis, we report that two additional pathways, including the SLT2 mitogen-activated protein kinase (MAPK) pathway and the osmosensing MAPK pathway, were also required for survival during ER stress. We further show that the SLT2 MAPK pathway was activated during ER stress, was responsible for increased resistance to ER stress, and functioned independently of the classic IRE1/HAC1 pathway. We propose that the SLT2 MAPK pathway is an important cell survival signaling pathway during ER stress. This study shows the feasibility of using the yeast deletion pool to identify relevant mammalian orthologues of the UPR.     

Sucrose regulated expression of a Ca2+-dependent protein kinase (TaCDPK1) gene in excised leaves of wheat.

Sucrose (Suc) can influence the expression of a large number of genes and thereby regulates many metabolic and developmental processes. However, the Suc sensing and the components of the ensuing signaling transduction pathway leading to the regulation of gene expression are not fully understood. We have shown that protein kinases and phosphatases are involved in the Suc induced expression of fructosyltransferase (FT) genes and fructan accumulation by an hexokinase independent pathway in wheat (Triticum aestivum). In the present study, using an RT-PCR based strategy, we have cloned a calcium-dependent protein kinase (TaCDPK1) cDNA that is upregulated during Suc treatment of excised wheat leaves. The deduced amino-acid sequence of CDPK1 has high sequence similarity (>70%) to known CDPKs from both monocots and dicots. Based on sequence homology, TaCDPK1 sequence shows a variable domain preceding a catalytic domain, an autoinhibitory function domain, and a C-terminal calmodulin-domain containing 4 EF-hand calcium-binding motifs, along with a N-myristoylation motif in the N-terminal variable domain. The recombinant Escherichia coli expressed TaCDPK1 was able to phosphorylate histone III-S in a calcium dependent manner in in vitro assays. The TaCDPK1 gene expression, as determined by quantitative RT-PCR, is induced by Suc and this effect is repressed by the inhibitors of the putative components of the Suc signal transduction pathway (calcium, Ser/Thr protein kinases and protein phosphatases). We propose that TaCDPK1 is involved in the Suc induced signaling pathway in wheat leaves.     

Regulation of Arabidopsis thaliana Ku genes at different developmental stages under heat stress

Ku, a heterodimeric protein consisting of 70- and 80-kDa subunits, is involved in many cellular processes, such as DNA replication, cell cycle regulation and heat shock response. Moreover, the expression of Arabidopsis thaliana Ku genes (AtKu) is modulated by certain plant hormones through several signal transduction pathways. This study investigated how AtKu are regulated by heat stress. AtKu expression in 3-week-old young seedlings was down-regulated by heat stress in a time-dependent manner, as examined using real-time quantitative PCR, GUS reporter systems, and western blotting analysis. Additionally, the heat-induced repression of AtKu was mediated through the abscisic acid (ABA) biosynthetic pathway, as shown by the reversal of AtKu suppression in the ABA biosynthesis mutant, aba3, and by an increase in the ABA level as analyzed by reverse-phase high performance liquid chromatography. Heat stress-induced regulation of AtKu repression also involved ethylene signaling, DNA repair pathways, and fatty acid synthesis. Furthermore, AtKu expression was repressed in stems, rosette leaves, and cauline leaves in 4-5-week-old plants under heat stress, whereas it remained unchanged in roots and primary inflorescence, indicating that heat differentially modulated AtKu expression in distinct tissues of Arabidopsis.     

Abscisic acid and jasmonic acid activate wound-inducible genes in potato through separate, organ-specific signal transduction pathways

Mechanical damage to leaf tissue causes an increase in abscisic acid (ABA) which in turn activates the biosynthesis of jasmonic acid (JA). The resulting higher endogenous JA levels subsequently activate the expression of wound-inducible genes. This study shows that JA induces the expression of different sets of genes in roots and leaves of potato plants. When roots of intact plants were treated with JA, high levels of proteinase inhibitor II (pin2), cathepsin D inhibitor, leucine aminopeptidase and threonine deaminase mRNAs accumulated in the systemic leaves. However, in the treated roots, very low, if any, expression of these genes could be detected. In contrast, a novel, root-specific pin2 homologue accumulated in the JA-treated root tissue which could not be detected in leaves, either systemic or those directly treated with JA. Application of okadaic acid and staurosporine revealed that a protein phosphorylation step is involved in the regulation of this differential response. In leaves, a protein phosphatase is required for the JA-induced expression of pin2 and the other genes analysed. This phosphatase activity is not necessary for the JA-induced expression of a pin2 homologue in roots, suggesting the existence of different transduction pathways for the JA signal in these organs. The requirement of a protein phosphatase activity for JA-mediated gene induction has enabled identification of a JA-independent pathway for ABA induction of pin2 and the other wound-inducible genes. This alternative pathway involves a protein kinase, and appears to be selective for wound-inducible genes. Our data suggest the presence of a complex, organ-specific transduction network for regulating the effects of the plant hormones ABA and JA on gene expression upon wounding.