Key features of heat shock regulatory elements.

Abstract

The promoters of heat shock protein genes are among the best-studied inducible eucaryotic promoters. Regions responsible for heat regulation have been identified previously by deletion experiments with several different heat shock genes. In this paper the critical importance of two novel features of heat shock regulatory elements was investigated. First, the elements were modular and, as a consequence, displayed a characteristic 5-nucleotide periodicity produced by multiple GAA blocks that were arranged in alternating orientations and at 2-nucleotide intervals. Functional heat shock regulatory elements appeared to include three or more of these blocks. Second, the nucleotides at the two positions immediately upstream from GAA segments played an important role in defining the competence of regulatory elements.Heat shock protein (hsp) genes occur in all cell types examined so far and are typically silent at the temperature of normal growth but are expressed at exceedingly high levels at elevated temperatures or in cells suffering from other types of stress (36). Drosophila melanogaster hsp70 genes encoding a major hsp of 70 kilodaltons (kDa) were the first hsp genes to be introduced into a variety of different cell types, such an NIH 3T3 cells (9), monkey COS cells (24, 30), Xenopus oocytes (3, 43), Drosophila cells (6,7,10,11,19,20), and others (see reference 27 for review). In most cell types, the genes were expressed in a heat-regulated fashion. A region located about 45 to 65 nucleotides upstream from the transcription start site of hsp70 genes was found to be essential for heat regulation in monkey cells and Xenopus oocytes (3,24,30). A second region containing related sequences, between about -65 and -90, was required in addition to the above region for high activity in Drosophila cells (1,11,39).Comparison of the -45 to -65 sequence of Drosophila hsp70 genes and of analogous sequences in other hsp genes led to the establishment of a heat shock consensus sequence, CNNGAANNTTCNNG, (31), where N is any nucleotide.Factors binding to this type of sequence have been identified (17,28,37,45,47,48), and two groups have purified such factors from Drosophila nuclear extracts and have shown that they are specifically involved in transcriptional activation of hsp genes (28,45,48).Since the sequences described above are the only elements specifically required for heat regulation and are also binding sites for hsp gene-specific transcription factors, they are of central importance for the understanding of heat shock regulation. Earlier studies in this laboratory (2) as well as by another group (40) had indicated that the heat shock consensus sequence that had been accepted for many years as the prototype heat shock regulatory sequence did not by itself function as a regulatory element. We have attempted here to define experimentally the nature of heat shock regulatory elements.MATERIALS AND METHODS Plasmid constructions. Plasmids ( Fig. 1 and 2) were derived from constructs D88 and D50 (2). In the latter constructs, D. m...

Mdj1p, a novel chaperone of the DnaJ family, is involved in mitochondrial biogenesis and protein folding

“…Upstream of the MDJ7 open reading frame, at positions -28 and -88 with respect to the translational start codon, are two potential TATA boxes. Furthermore, the MDJ7 open reading frame is preceded by several weak matches to the heat shock element nGAAnnlXn (Amin et al, 1988;Young and Craig, 1993) and five copies of the CCCCT motif reported to be involved in heat shockfactor-independent gene induction (Kobayashi and McEntee, 1993) (Figure 1). The heat inducibility of MDJl was tested by Northern blot analysis, which revealed a 2-to 3-fold transient increase of transcript abundance upon shift of the cells from 24% to 37% (data not shown).…”

Hsp70 Inhibits Aminoglycoside-Induced Hair Cell Death and is Necessary for the Protective Effect of Heat Shock

“…Stressinduced activation of Hsf1 involves trimerization, acquisition of DNA-binding activity, and translocation to the nucleus (Baler et al 1993;Sarge et al 1993). In the nucleus, Hsf1 binds to heat shock element (HSE) sequences present in the promoter regions of Hsps (Amin et al 1988). In addition, regulation of Hsf1-mediated transcriptional activity is mediated by both constitutive and stress-inducible phosphorylation (Kline and Morimoto 1997;Holmberg et al 2001Holmberg et al , 2002Guettouche et al 2005).…”

Insights into electromagnetic interaction mechanisms

“…The HSEs for heatresponsiveness are located in the 5 0 -flanking sequences of the HSP70 genes that contain triplet repeats of nGAAn, a nucleotide recognition motif/sequence (Pelham, 1982). A minimum of three 5-bp modules are required to form a functional HSE (Amin et al, 1988). The heat shock domain on the HSP70 promoter lies between À 107 and À 68, relative to the transcription initiation site, with the HSE centered at À 100.…”