Stress-induced Transcription Factor Research Articles

Liver gene expression analysis reveals endoplasmic reticulum stress and metabolic dysfunction in SCD1-deficient mice fed a very low-fat diet.

We previously reported that mice deficient in stearoyl-CoA desaturase-1 (Scd1) and maintained on a very low-fat (VLF) diet for 10 days developed severe loss of body weight, hypoglycemia, hypercholesterolemia, and many cholestasis-like phenotypes. To better understand the metabolic changes associated with these phenotypes, we performed microarray analysis of hepatic gene expression in chow- and VLF-fed female Scd1+/+ and Scd1-/- mice. We identified an extraordinary number of differentially expressed genes (>4,000 probe sets) in the VLF Scd1-/- relative to both VLF Scd1+/+ and chow Scd1-/- mice. Transcript levels were reduced for genes involved in detoxification and several facets of fatty acid metabolism including biosynthesis, elongation, desaturation, oxidation, transport, and ketogenesis. This pattern is attributable to the decreased mRNA abundance of several genes encoding key transcription factors, including LXRalpha, RXRalpha, FXR, PPARalpha, PGC-1beta, SREBP1c, ChREBP, CAR, DBP, TEF, and HLF. A robust induction of endoplasmic reticulum (ER) stress is indicated by enhanced splicing of XBP1, increased expression of the stress-induced transcription factors CHOP and ATF3, and elevated expression of several genes involved in the integrated stress and unfolded protein response pathways. The gene expression profile is also consistent with induction of an acute inflammatory response and macrophage recruitment. These results highlight the importance of monounsaturated fatty acid synthesis for maintaining metabolic homeostasis in the absence of sufficient dietary unsaturated fat and point to a novel cellular nutrient-sensing mechanism linking fatty acid availability and/or composition to the ER stress response.

Regulation of Thermotolerance by Stress-Induced Transcription Factors in Saccharomyces cerevisiae

The heat shock transcription factor Hsf1 and the general stress transcription factors Msn2 and Msn4 (Msn2/4) are major regulators of the heat shock response in Saccharomyces cerevisiae. Here, we show that transcriptional activation of their target genes, including HSP104, an antistress chaperone gene, is obligatory for thermotolerance. Although Hsf1 activity might be necessary before the exposure of cells to high temperature, severe heat shock induced the binding of hyperphosphorylated Hsf1 to its target promoters. However, promoter-bound, phosphorylated Hsf1 was inactive for transcription because RNA polymerase II was inactive at high temperatures. Rather, our results suggest that Hsf1 activates the transcription of most of its target genes during the recovery period following severe heat shock. This delayed upregulation by Hsf1, which would be induced by misfolded proteins that accumulate in severely heat-shocked cells, is required for the resumption of normal cell growth. In contrast, the factors Msn2/4 were not involved in the delayed upregulation of genes and were dispensable for cell growth during the recovery period, suggesting that they play a role before the exposure to high temperature. These results show that Hsf1 and Msn2/4 act differentially before and after exposure to extreme temperatures to ensure cell survival and growth.

Acute hypoxia to endothelial cells induces activating transcription factor 3 (ATF3) expression that is mediated via nitric oxide

Endothelial cells (ECs) play an important role in hypoxia-induced vascular disorders. We investigated the acute hypoxia effect on endothelial expression of activating transcription factor 3 (ATF3), a stress-inducible transcription factor playing significant roles in cellular responses to stress. Bovine aortic ECs were subjected to acute hypoxia (1% O 2, pO 2 = 8 mmHg) and ATF3 expression was examined. ECs exposed to hypoxia transiently induced ATF3 expression. A transient increase in the activation of c-Jun-NH 2-terminal kinase (JNK) and p38 mitogen-activated protein kinase (MAPK) in ECs was observed; however, only ECs pretreated with a specific inhibitor to JNK suppressed the hypoxia-induced ATF3 expression. ECs exposed to acute hypoxia transiently increased endothelial nitric oxide (eNOS) activity. Pre-treating ECs with a specific inhibitor to eNOS ( l-NAME) or PI3-kinase significantly inhibited the hypoxia-induced JNK activation and ATF3 expression. ATF3 induction has been shown to inhibit matrix metalloproteinase-2 (MMP-2) expression. Consistently, ECs exposed to hypoxia attenuated the MMP-2 expression. This hypoxia-attenuated MMP-2 expression can be rescued by pre-treating ECs with an inhibitor of eNOS. These results suggest that the ATF3 induction by acute hypoxia is mediated by nitric oxide and the JNK pathway in ECs. Our findings provide a molecular basis for the mechanism in which ECs respond to acute hypoxia.

Scd1 deficiency causes endoplasmic reticulum stress and metabolic dysfunction in mice fed a very low‐fat diet

We performed microarray analysis of hepatic gene expression in mice deficient in stearoyl‐CoA desaturase‐1 (Scd1) and maintained on a chow or a very low‐fat (VLF) diet for 10 days. We identified an extraordinary number of differentially expressed genes (>4000 probe sets) in the VLF Scd1−/− relative to both VLF Scd1+/+ and chow Scd1−/− mice. Transcript levels were reduced for genes involved in several facets of fatty acid metabolism including biosynthesis, elongation, desaturation, oxidation, transport, and ketogenesis. This pattern is attributable to the decreased mRNA abundance of several key lipid metabolism transcription factors, including LXRα, RXRα, PPARα, PGC‐1β, SREBP1c, and ChREBP. A robust induction of endoplasmic reticulum (ER) stress is indicated by the increased expression of stress‐induced transcription factors, including CHOP and ATF3, and several of their target mRNAs. Activation of the unfolded protein response pathway is further evidenced by increased splicing of XBP1 mRNA. The gene expression profile is also consistent with induction of an acute inflammatory response and macrophage recruitment. These results highlight the importance of monounsaturated fatty acid synthesis for maintaining metabolic homeostasis in the absence of sufficient dietary unsaturated fat and point to a novel cellular nutrient‐sensing mechanism linking fatty acid availability and/or composition to the ER stress response. Supported by NIH grants HL56593, DK58037, and DK066369.

Impaired Cytoprotective Mechanisms in Eyes with Pseudoexfoliation Syndrome/Glaucoma

Evidence suggests that chronically increased stress conditions in the anterior eye segment constitute major mechanisms involved in the pathobiology of pseudoexfoliation (PEX) syndrome. The expression of stress-related genes in eyes from patients with and without PEX syndrome/glaucoma was investigated to determine whether PEX syndrome is associated with an altered cellular stress response. cDNA array hybridization, quantitative real-time PCR, Western blot analysis, and immunohistochemistry were applied to analyze the mRNA and protein expression of stress-related genes in anterior segment tissues of PEX eyes, with and without glaucoma, and to compare them with normal and glaucomatous control eyes. Hybridization of cDNA arrays identified a set of stress-related candidate genes for differential expression in PEX syndrome/glaucoma, of which 10 were confirmed by real-time PCR in ciliary processes and iris tissue. The expression of MAPKp38, heat shock proteins (HSP40, HSP60), and superoxide dismutase (SOD2) was increased up to threefold in PEX specimens. In contrast, a large set of cytoprotective gene products, including antioxidant defense enzymes (glutathione S-transferases mGST1 and GSTT1), ubiquitin-conjugating enzymes (UBE2A, UBE2B), the DNA repair protein MLH1, and the stress-inducible transcription factor GADD153, were found to be consistently downregulated up to threefold in PEX specimens on both the mRNA and protein levels. The present findings provide evidence of alterations in cytoprotective mechanisms including antioxidant defense, proteasome function, endoplasmic reticulum-related stress response, and DNA repair in anterior segment tissues of PEX eyes. The resultant enhanced sensitivity and vulnerability to cellular stress conditions may therefore be one contributing factor in the pathobiology of PEX syndrome.

Identification of stress-tolerance-related transcription-factor genes via mini-scale Full-length cDNA Over-eXpressor (FOX) gene hunting system

Recently, we developed a novel system known as Full-length cDNA Over-eXpressor (FOX) gene hunting [T. Ichikawa, M. Nakazawa, M. Kawashima, H. Iizumi, H. Kuroda, Y. Kondou, Y. Tsuhara, K. Suzuki, A. Ishikawa, M. Seki, M. Fujita, R. Motohashi, N. Nagata, T. Takagi, K. Shinozaki, M. Matsui, The FOX hunting system: an alternative gain-of-function gene hunting technique, Plant J. 48 (2006) 974–985], which involves the random overexpression of a normalized Arabidopsis full-length cDNA library. While our system allows large-scale collection of full-length cDNAs for gene discovery, we sought to downsize it to analyze a small pool of full-length cDNAs. As a model system, we focused on stress-inducible transcription factors. The full-length cDNAs of 43 stress-inducible transcription factors were mixed to create a transgenic plant library. We screened for salt-stress-resistant lines in the T1 generation and identified a number of salt-tolerant lines that harbored the same transgene ( F39). F39 encodes a bZIP-type transcription factor that is identical to AtbZIP60, which is believed to be involved in the endoplasmic reticulum stress response. Microarray analysis revealed that a number of stress-inducible genes were up-regulated in the F39-overexpressing lines, suggesting that AtbZIP60 is involved in stress signal transduction. Thus, our mini-scale FOX system may be used to screen for genes with valuable functions, such as transcription factors, from a small pool of genes that show similar expression profiles.

ER Stress Triggers Apoptosis by Activating BH3-Only Protein Bim

Endoplasmic reticulum (ER) stress caused by misfolded proteins or cytotoxic drugs can kill cells and although activation of this pathway has been implicated in the etiology of certain degenerative disorders its mechanism remains unresolved. Bim, a proapoptotic BH3-only member of the Bcl-2 family is required for initiation of apoptosis induced by cytokine deprivation or certain stress stimuli. Its proapoptotic activity can be regulated by several transcriptional or posttranslational mechanisms, such as ERK-mediated phosphorylation, promoting its ubiquitination and proteasomal degradation. We found that Bim is essential for ER stress-induced apoptosis in a diverse range of cell types both in culture and within the whole animal. ER stress activates Bim through two novel pathways, involving protein phosphatase 2A-mediated dephosphorylation, which prevents its ubiquitination and proteasomal degradation and CHOP-C/EBPalpha-mediated direct transcriptional induction. These results define the molecular mechanisms of ER stress-induced apoptosis and identify targets for therapeutic intervention in ER stress-related diseases.

S‐Adenosylmethionine Attenuates Hepatic Lipid Synthesis in Micropigs Fed Ethanol With a Folate‐Deficient Diet

To demonstrate a causative role of abnormal methionine metabolism in the pathogenesis of alcoholic steatosis, we measured the effects on hepatic lipid synthesis of supplementing ethanol and folate-deficient diets with S-adenosylmethionine (SAM), a metabolite that regulates methionine metabolism. Yucatan micropigs were fed folate-deficient diets as control, with ethanol at 40% of kcal, and with ethanol supplemented with SAM at 0.4 g/1,000 kcal for 14 weeks. Histopathology, triglyceride levels and transcripts, and protein levels of the regulatory signals of hepatic lipid synthesis were measured in terminal omental adipose and liver samples. Feeding ethanol at 40% of kcal with folate-deficient diets for 14 weeks increased and supplemental SAM maintained control levels of liver and plasma triglyceride. Serum adiponectin, liver transcripts of adiponectin receptor-1 (AdipoR1), and phosphorylated adenosine monophosphate kinase-beta (p-AMPKbeta) were each reduced by ethanol feeding and were sustained at normal levels by SAM supplementation of the ethanol diets. Ethanol feeding activated and SAM supplementation maintained control levels of ER stress-induced transcription factor sterol regulatory element-binding protein-1c (SREBP-1c) and its targeted transcripts of lipid synthesizing enzymes acetyl-CoA carboxylase (ACC), fatty acid synthase (FAS), and glycerol-3-phosphate acyltransferase (GPAT). Ethanol feeding with a folate-deficient diet stimulates hepatic lipid synthesis by down-regulating adiponectin-mediated pathways of p-AMPK to increase the expression of nSREBP-1c and its targeted lipogenic enzymes. Preventing abnormal hepatic methionine metabolism by supplementing ethanol diets with SAM reduces liver triglyceride levels by up-regulation of adiponectin-mediated pathways to decrease fatty acid and triglyceride synthesis. This study demonstrates that ethanol-induced hepatic lipid synthesis is mediated in part by abnormal methionine metabolism, and strengthens the concept that altered methionine metabolism plays an integral role in the pathogenesis of steatosis.

HSF1 is necessary for reactive oxygen species (ROS) homeostasis and proliferation in T cells at fever temperatures. (87.47)

Abstract HSF1 is a major heat and stress inducible transcription factor in eukaryotic cells responsible for the regulation of a variety of genes involved in maintenance of cellular homeostasis. We have previously shown that HSF1 is activated at physiologic fever temperatures (39oC) in T cells vs. much higher, non-physiologic temperatures (42oC) in most other cell types. T cells from HSF1−/− mice proliferate normally at 37oC, but are severely inhibited at 39oC. These T cells are blocked at the G1-S phase transition of the cell cycle. T cells normally increase ROS levels as a result of activation. Antioxidants which block ROS increases also inhibit proliferation. We found that fever temperatures also inhibited the normal activation induced increase of ROS in T cells within 2h. Wild type T cells can overcome this inhibition in ROS generation within 5h and go on to proliferate normally. In contrast, HSF1−/− T cells were unable to overcome this dysregulation and failed to proliferate. Thus, fever temperature causes a dysregulation in ROS homeostasis in T cells and HSF1 is critical in restoring this regulation.

Hypoxic Regulation of Id-1 and Activation of the Unfolded Protein Response Are Aberrant in Neuroblastoma

The Id proteins play an important role in proliferation, differentiation and tumorigenesis. Many tumors are hypoxic, but it is unknown if expression of Id proteins is regulated in hypoxic cells. Here we show that Id-1 is down-regulated in multiple primary, immortalized, and neoplastic hypoxic cell lines, and the transcriptional repressor ATF-3 is both necessary and sufficient for this hypoxia-induced repression of Id-1. Hypoxic up-regulation of ATF-3 is due in part to activation of the unfolded protein response, a cellular stress response. Remarkably, we observe that the unfolded protein response is de-regulated in all neuroblastoma cell lines tested. Indeed, in the absence of ATF-3 the hypoxia-induced transcription factor HIF-1 up-regulates Id-1 in hypoxic neuroblastoma cells. Hypoxic neuroblastoma cells diminish expression of some neuronal differentiation markers, and forced expression of ATF-3 in hypoxic neuroblastoma cells represses Id-1 and prevents the loss of these markers. The divergent regulation of Id proteins in distinct hypoxic cells may explain some of the varied effects hypoxia has on cellular differentiation and proliferation.

Endoplasmic Reticulum Stress Is a Target for Therapy in Waldenstrom's Macroglobulinemia (WM).

Background: WM is an incurable low-grade lymphoplasmacytic lymphoma with limited options of therapy. Proteasome inhibition has been shown to induce components of the proapoptotic/terminal unfolded protein response (UPR), a signaling pathway activated by accumulation of misfolded proteins within the endoplasmic reticulum (ER). We previously found that UPR gene expression is related to disease activity in WM with a particular role for GRP78/Bip as a prognostic factor. We therefore examined tunicamycin (Sigma, St Louis, MO), a potent inducer of ER stress, for potential anti-tumor effects in WM.Methods: WM cell lines (BCWM.1 and WSU-WM), IgM secreting low-grade lymphoma cell lines (MEC1, RL) and primary CD19+ selected LPC cells from WM patients were incubated with tunicamycin (0.01–10 uM) for 24–72 hours and evaluated by MTT, thymidine uptake, and Apo2.7/PI staining for effects on proliferation and survival. Since bone marrow stromal cells (BMSC) confer growth and resistance to conventional treatments, we also tested the effect of tunicamycin on WM cells co-cultured with BMSC. Immunoblotting for caspases was also performed and expression of UPR genes determined using relative quantitative RT-PCR reaction following (0.5–16 hrs) culture with tunicamycin.Results: WM cells inherently expressed the ER chaperones GRP78/Bip and GRP94/gp96. Tunicamycin rapidly induced components of the proapoptotic/terminal UPR, including PERK, the ER stress-specific eIF-2alpha kinase; ATF6, an ER stress-induced transcription factor; and its proapoptotic target, CHOP/GADD153. Tunicamycin also induced significant cytotoxicity, and inhibited DNA synthesis with an IC50 of 0.5–1 ug/mL in all cell lines, as well as primary LPC from 3/3 WM patients. Furthermore, tunicamycin induced apoptosis in WM cells, with an increase in the sub-G1 population notable at 12 hrs. Tunicamycin induced apoptosis was preceded by caspase-12 cleavage, followed then by caspase-8, -9 and PARP cleavage. Importantly, co-culture of WM cells with the survival factors IL-6, IGF-1 as well as BMSC did not inhibit tunicamycin induced cytotoxicity. Lastly, tunicamycin did not induce cytotoxicity in healthy donor peripheral blood mononuclear or hematopoietic stem cells.Conclusion: These pre-clinical studies provide a framework for further evaluation of ER stress inducing agents as therapeutic agents in WM.

Stress-inducible DREB2A transcription factor from Pennisetum glaucum is a phosphoprotein and its phosphorylation negatively regulates its DNA-binding activity

Abiotic stress-mediated gene expression is regulated via different transcription factors of which drought-responsive element-binding (DREB) proteins play an important role. There are two types of DREBs. Presently, the function of DREB1 type protein is well studied; however, much less information is available for DREB2. In this study, a cDNA with an open reading frame of 332 amino acids, encoding the transcription activation factor DREB2A, was cloned from Pennisetum glaucum, a stress tolerant food grain crop. Phylogenetic tree revealed that PgDREB2A is more close to DREBs isolated from monocots, though it forms an independent branch. The PgDREB2A transcript was up-regulated in response to drought within 1 h of the treatment, whereas the induction was delayed in response to cold and salinity stress. However, during cold stress, the transcript was induced more as compared to drought and salinity. The recombinant PgDREB2A protein having a molecular mass of 36.6 kDa was purified using Ni-NTA affinity chromatography. Gel mobility shift assays using the purified protein and two cis elements of rd29A (responsive to dehydration 29A) gene promoter of Arabidopsis revealed that PgDREB2A binds to drought-responsive element (DRE) ACCGAC and not to GCCGAC. PgDREB2A is a phosphoprotein, which has not been reported earlier. The phosphorylation of PgDREB2A in vitro by P. glaucum total cell extract occurred at threonine residue(s). The phosphorylated PgDREB2A did not bind to the DREs. The present data indicate that stress induction of genes could occur via post-translational modification by phosphorylation of DREB2A.

Inhibitory effect of 4-(2-aminoethyl)-benzenesulfonyl fluoride, a serine protease inhibitor, on PI3K inhibitor-induced CHOP expression

Endoplasmic reticulum stress contributes to several diseases such as neurodegenerative disorders and diabetes. In the previous report, we found that phosphatidylinositol 3-kinase (PI3K) down-regulation is important for inducing CHOP expression, an endoplasmic reticulum stress-induced transcription factor. In the present study, we investigated the effect of 4-(2-aminoethyl)-benzenesulfonyl fluoride (AEBSF), a serine protease inhibitor, on PI3K inhibitor-induced CHOP expression. We found that AEBSF completely inhibited PI3K inhibitor-induced CHOP expression at both mRNA and protein levels. It is suggested that AEBSF is an important drug from a pharmacological point of view and the results may have important implications for understanding endoplasmic reticulum stress-related diseases.

Proteasome inhibitors induce a terminal unfolded protein response in multiple myeloma cells

AbstractMultiple myeloma (MM) is an incurable plasma cell malignancy. The 26S proteasome inhibitor, bortezomib, selectively induces apoptosis in MM cells; however, the nature of its selectivity remains unknown. Here we demonstrate that 5 different MM cell lines display similar patterns of sensitivity to 3 proteasome inhibitors (PIs) but respond differently to specific NF-κB inhibition. We further show that PIs initiate the unfolded protein response (UPR), a signaling pathway activated by the accumulation of misfolded proteins within the endoplasmic reticulum (ER). Consistent with reports that prosurvival/physiologic UPR components are required for B-cell differentiation into antibody-secreting cells, we found that MM cells inherently expressed the ER chaperones GRP78/Bip and GRP94/gp96. However, bortezomib rapidly induced components of the proapoptotic/terminal UPR, including PERK, the ER stress–specific eIF-2α kinase; ATF4, an ER stress–induced transcription factor; and its proapoptotic target, CHOP/GADD153. Consistent with our hypothesis that PIs induce the accumulation of misfolded ER-processed proteins, we found that the amount of immunoglobulin subunits retained within MM cells correlated with their sensitivity to PIs. These findings suggest that MM cells have a lower threshold for PI-induced UPR induction and ER stress–induced apoptosis because they constitutively express ER stress survival factors to function as secretory cells.

Selective Inhibition of Endoplasmic Reticulum-associated Degradation Rescues ΔF508-Cystic Fibrosis Transmembrane Regulator and Suppresses Interleukin-8 Levels: THERAPEUTIC IMPLICATIONS

Selective Inhibition of Endoplasmic Reticulum-associated Degradation Rescues ΔF508-Cystic Fibrosis Transmembrane Regulator and Suppresses Interleukin-8 Levels: THERAPEUTIC IMPLICATIONS

Lipoic acid supplementation prevents cyclosporine-induced hypertension and nephrotoxicity in spontaneously hypertensive rats

Cyclosporine (CsA) has significantly improved long-term survival after organ transplantations. Hypertension and nephrotoxicity are common side effects during CsA treatment and are aggravated by high salt intake. To examine whether lipoic acid (LA), a natural antioxidant that scavenges reactive oxygen species and regenerates/recycles endogenous antioxidants, could prevent CsA-induced hypertension and nephrotoxicity. Six-week-old spontaneously hypertensive rats (SHR) on a high-sodium diet (NaCl 6%) received CsA [5 mg/kg subcutaneously (s.c.)] alone or in combination with LA (0.5% w/w) for 6 weeks. Blood pressure, arterial functions, and tissue morphology were determined. Immunohistochemistry, quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) and high-pressure liquid chromatography were used for kidney and heart samples. CsA induced severe hypertension, cardiac hypertrophy, endothelial dysfunction, and pronounced albuminuria. Histologically, the kidneys showed severe thickening of the media of the afferent arteries with fibrinoid necrosis, perivascular monocyte/macrophage infiltration and nitrotyrosine overexpression. CsA induced the expression of fibrogenic connective tissue growth factor both in the heart and kidneys. The detrimental effects of CsA were associated with upregulation of myocardial atrial natriuretic peptide (ANP) mRNA expression, paradoxical activation of the renin-angiotensin system (RAS), induction of renal reduced nicotinamide-adenine dinucleotide phosphate (NADPH) oxidase, and overexpression of oxidative stress-induced transcription factor NRF2. LA lowered blood pressure, ameliorated cardiac hypertrophy and endothelial dysfunction, and totally normalized albuminuria. In LA-treated rats, renal and cardiac morphologies were indistinguishable from those of SHR controls. CsA-induced myocardial ANP and connective tissue growth factor (CTGF) mRNA overexpression, RAS activation, NADPH oxidase induction, and NRF2 overexpression were prevented by LA. LA induced the mRNA expression of gamma-glutamylcysteine ligase, the rate-limiting enzyme in glutathione synthesis, and markedly increased hepatic cysteine and glutathione concentrations. Our findings suggest a salutary role for lipoic acid supplementation in the prevention of CsA-induced hypertension and nephrotoxicity, and underscore the importance of increased oxidative stress in the pathogenesis of CsA toxicity.

低温および乾燥環境下におけるシロイヌナズナのトランスクリプト-ムおよびメタボロ-ム解析

低温および乾燥環境下におけるシロイヌナズナのトランスクリプト-ムおよびメタボロ-ム解析

Expression of the Arabidopsis DREB1A gene in transgenic chrysanthemum enhances tolerance to low temperature

SummaryThe gene DREB1A, encoding a stress-inducible transcription factor, driven by the cauliflower mosaic virus 35S (CaMV) promoter or by the stress-inducible rd29A promoter, was transferred into chrysanthemum plants [Dendranthema grandiflorum (Ramat.) Kitamura] cv. ‘Fall Color’ by Agrobacterium-mediated transformation. Integration of the target gene, DREB1A was confirmed by PCR and Southern blotting. When exposed to 2°C, expression of DREB1A was enhanced in the roots of young transgenic 35S:DREB1A plant lines, and induced in transgenic rd29A:DREB1A lines. Electrolyte leakage in leaves of rd29A:DREB1A plant lines was significantly lower than in 35S:DREB1A lines. Compared to control plants, superoxide dismutase activities and proline contents increased slowly in transgenic plants at the start of the cold stress treatment and remained at high levels during later periods, especially in rd29A:DREB1A transgenic lines. Young plants of the rd29A:DREB1A line could tolerate –8°C for 12 h, with a survival rate of 37.5%. No survival was observed in 35S:DREB1A lines or in wild-type plants. These results indicate that a combination of the stress-inducible rd29A promoter and the DREB1A gene enhanced the tolerance of ground-cover chrysanthemum plants to cold stress through a transgenic approach.

PI3K-Akt inactivation induced CHOP expression in endoplasmic reticulum-stressed cells

Stress signals that impair the function of the endoplasmic reticulum (ER) can lead to an accumulation of unfolded proteins in the ER causing cell death. Recent studies have indicated that ER stress contributes to several diseases such as neurodegenerative disorders or diabetes. In the present study, we found that Akt down-regulation is important for inducing CHOP expression, an ER stress-induced transcription factor. Treatment with tunicamycin or thapsigargin, ER stress inducers, caused dephosphorylation of Akt from 12 to 24 h and induced cell death. Interestingly, treatment with a PI3K inhibitor alone induced CHOP expression and caused cell death. However, a MEK1 inhibitor induced neither CHOP expression nor cell death. These results indicate that the inactivation of Akt by ER stress induces CHOP expression and causes cell death. Therefore, Akt plays an important role in ER stressed condition and may have important implications for understanding ER stress-related diseases.

Improvement of wheat drought and salt tolerance by expression of a stress-inducible transcription factorGmDREB of soybean (Glycine max)

Under stress conditions such as drought, high-salinity and low-temperature, the transcription factor of DREB (dehydration responsive element binding proteins) improved efficiently stress resistance by regulating the expression of its downstream genes with various environmental stress resistance in plants.GmDREB gene (GenBank Accession No. AF514908) encoding a stress-inducible transcription factor was cloned by screening a cDNA library ofGlycine max cv. Jinong 27 with yeast one-hybrid method.GmDREB gene was 910 bp in length and encoded 174 amino acids containing a conserved AP2/EREBP DNA-binding domain of 58 amino acids. Two conserved functional amino acids, valine and glutamic acid, were located on the 14th and the 19th amino acid residues in the conserved structural domain. An alkaline amino acid region (KKR) related to a nuclear localization signal was at the N-terminal, while an acidic amino acid region (DDD) related totrans-activation was at the C-terminal. Plant expression vectors were constructed and transformed into wheat by bombardment. In total, 13 transgenic plants withUbi::GmDREB and 11 transgenic plants withrd29A::GmDREB were identified from 103 regeneration plants by molecular analysis. The drought and salt tolerances of T1 transgenic lines withUbi::GmDREB orrd29A::GmDREB were demonstrated to be improved as compared to wild type. The result also suggested that bothUbiquitin andrd29A promoters could effectively drive the expression of theGmDREB gene and enhance drought and salt tolerance of T1 plants.