Sodium Salicylate Decreases Intracellular ATP, Induces Both Heat Shock Factor Binding and Chromosomal Puffing, but Does Not Induce hsp 70 Gene Transcription in Drosophila

Neil A Winegarden,Ken S Wong,Mary Sopta,J Timothy Westwood

doi:10.1074/jbc.271.43.26971

Abstract

Sodium salicylate has long been known to be an inducer of the heat shock puffs and presumably heat shock gene transcription in the polytene chromosomes of Drosophila salivary gland cells. Stress-induced transcription of the heat shock genes is mediated by the transcription factor known as Heat Shock Factor (HSF). In yeast, sodium salicylate has been reported to induce the DNA binding of HSF but not heat shock gene transcription itself, and similar findings have been reported in human cells. This apparent discrepancy in the induction of certain aspects of the heat shock response between these organisms prompted us to carefully reexamine the induction of the heat shock response in Drosophila salivary gland cells of third instar larvae and Drosophila tissue culture (SL2) cells. Sodium salicylate (3-30 mM) decreases intracellular ATP levels in SL2 cells and induces HSF binding activity in SL2 and salivary gland cells in a dose-dependent manner. Despite the induction of HSF binding and heat shock puffs in polytene chromosomes, we found no evidence for increased hsp 70 gene transcription suggesting that chromosomal puffing and gene transcription may be separable events. Salicylate did not induce the HSF hyperphosphorylation that is normally associated with HSF activation. Furthermore, salicylate (30 mM) prevented heat-induced hyperphosphorylation of HSF and hsp 70 gene transcription indicating that salicylate's inhibitory effect on hsp 70 transcription may be independent of its effect on HSF binding activity. We propose that the reduction in intracellular ATP caused by the addition of salicylate likely plays a role in the activation of HSF binding and the inhibition of both HSF hyperphosphorylation and hsp 70 gene transcription.

Highlights

One of the most widely administered family of drugs is the anti-inflammatory salicylates
Salicylate Induces heat shock factor (HSF) Binding and Chromosomal Puffing at Heat Shock Gene Loci in Polytene Chromosomes of D. melanogaster Salivary Glands—HSF has been shown previously to undergo a change in its DNA binding pattern on the polytene chromosomes of D. melanogaster third instar salivary gland cells when the larvae are exposed to heat stress [22]
It was observed in the present series of experiments that the previously published in vivo findings could be duplicated in an in vitro heat shock assay (Fig. 1B)

Summary

Salicylate Induces a Partial Heat Shock Response in Drosophila

Salicylate treatments (3–30 mM) do not induce transcription of the hsp 70 heat shock genes in salivary glands nor in SL2 cells as assayed by primer extension analysis. These results suggest that chromosomal puffing and gene transcription are likely separable events. Similar to what is seen in yeast, salicylate (30 mM) prevents heatinduced transcription of the heat shock genes It is not known how salicylate activates HSF binding, but models have been proposed which indicate that the lowering of intracellular pH [19] or the accumulation of newly synthesized aberrant proteins [17, 18] might be involved in the activation process. A model as to how lowered ATP levels might activate HSF binding and prevent transcription of the heat shock genes is discussed

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION