Synergistic effects of HSE and LTR elements from hsp70 gene promoter of Ulva prolifera (Ulvophyceae,Chlorophyta) upon temperature induction1

Besides heat stress, the 70 kDa heat shock proteins (HSP70s) have been shown to respond to cold stress. However, the involved cis‐acting elements remain unknown. The hsp70 gene from the green macroalga Ulva prolifera (Uphsp70) has been cloned, from which one heat shock element HSE and one low‐temperature‐responsive element LTR were found in the promoter. Using the established transient expression system and quantitative GUS assay, a series of element deletion experiments were performed to determine the functions of HSE and LTR in response to temperature stress. The results showed that under cold stress, both HSE and LTR were indispensable, since deletion leads to complete loss of promoter activity. Under heat stress, although the HSE could respond independently, coexistence with LTR was essential for high induced activity of the Uphsp70 promoter. Therefore, synergistic effects exist between HSE and LTR elements in response to temperature stress in Ulva, and extensive bioinformatics analysis showed that the mechanism is widespread in algae and plants, since LTR coexists widely with HSE in the promoter region of hsp70. Our findings provide important supplements to the knowledge of algal and plant HSP70s response to temperature stress. We speculated that for algal domestication and artificial breeding, HSE and LTR elements might serve as potential molecular targets to temperature acclimation.     

Heat stress-induced H2O2 is required for effective expression of heat shock genes in Arabidopsis

The mechanisms of sensing and signalling of heat and oxidative stresses are not well understood. The central question of this paper is whether in plant cells oxidative stress, in particular H₂O₂, is required for heat stress- and heat shock factor (HSF)-dependent expression of genes. Heat stress increases intracellular accumulation of H₂O₂ in Arabidopsis cell culture. The accumulation was greatly diminished using ascorbate as a scavenger or respectively diphenyleneiodonium chloride (DPI) as an inhibitor of reactive oxygen species production. The mRNA of heat shock protein (HSP) genes, exemplified by Hsp17.6, Hsp18.2, and the two cytosolic ascorbate peroxidase genes Apx1, Apx2, reached similar levels by moderate heat stress (37°C) or by treatment with H₂O₂, butylperoxide and diamide at room temperature. The heat-induced expression levels were significantly reduced in the presence of ascorbate or DPI indicating that H₂O₂ is an essential component in the heat stress signalling pathway. Rapid (15 min) formation of heat shock promoter element (HSE) protein-binding complex of high molecular weight in extracts of heat-stressed or H₂O₂-treated cells and the inability to form this complex after ascorbate treatment suggests that oxidative stress affects gene expression via HSF activation and conversely, that H₂O₂ is involved in HSF activation during the early phase of heat stress. The heat stress induction of a high mobility HSE-binding complex, characteristic for later phase of heat shock response, was blocked by ascorbate and DPI. H₂O₂ was unable to induce this complex suggesting that H₂O₂ is involved only in the early stages of HSF activation. Significant induction of the genes tested after diamid treatment and moderate expression of the sHSP genes in the presence of 50 mM ascorbate at 37°C occurred without activation of HSF, indicating that other mechanisms may be involved in stress signalling. Electronic Supplementary Material Supplementary material is available for this article at http//dx.doi.org/10.1007/s11103-006-0045-4 Roman A. Volkov and Irina I. Panchuk contributed equally     

Geldanamycin restores a defective heat shock response in vivo

Induced expression of heat shock proteins (Hsps) plays a central role in promoting cellular survival after environmental and physiological stress. We have previously shown that scrapie-infected mouse neuroblastoma (ScN2a) cells fail to induce the expression of Hsp72 and Hsp28 after various stress conditions. Here we present evidence that this impaired stress response is due to an altered regulation of HSF1 activity. Upon stress in ScN2a cells, HSF1 was converted into hyperphosphorylated trimers but failed to acquire transactivation competence. A kinetic analysis of HSF1 activation revealed that in ScN2a cells trimer formation after stress was efficient, but disassembly of trimers proceeded much faster than in the uninfected cell line. Geldanamycin, a Hsp90-binding drug, significantly delayed disassembly of HSF1 trimers after a heat shock and restored stress-induced expression of Hsp72 in ScN2a cells. Heat-induced Hsp72 expression required geldanamycin to be present; following removal of the drug ScN2a cells again lost their ability to mount a stress response. Thus, our studies show that a defective stress response can be pharmacologically restored and suggest that the HSF1 deactivation pathway may play an important role in the regulation of Hsp expression.