The DNA-Binding Properties of Two Heat Shock Factors, HSF1 and HSF3, Are Induced in the Avian Erythroblast Cell Line HD6

Abstract

Avian cells express three heat shock transcription factor (HSF) genes corresponding to a novel factor, HSF3, and homologs of mouse and human HSF1 and HSF2. Analysis of the biochemical and cell biological properties of these HSFs reveals that HSF3 has properties in common with both HSF1 and HSF2 and yet has features which are distinct from both. HSF3 is constitutively expressed in the erythroblast cell line HD6, the lymphoblast cell line MSB, and embryo fibroblasts, and yet its DNA-binding activity is induced only upon exposure of HD6 cells to heat shock. Acquisition of HSF3 DNA-binding activity in HD6 cells is accompanied by oligomerization from a non-DNA-binding dimer to a DNA-binding trimer, whereas the effect of heat shock on HSF1 is oligomerization of an inert monomer to a DNA-binding trimer. Induction of HSF3 DNA-binding activity is delayed compared with that of HSF1. As occurs for HSF1, heat shock leads to the translocation of HSF3 to the nucleus. HSF3 exhibits the properties of a transcriptional activator, as judged from the stimulatory activity of transiently overexpressed HSF3 measured by using a heat shock element-containing reporter construct and as independently assayed by the activity of a chimeric GAL4-HSF3 protein on a GAL4 reporter construct. These results reveal that HSF3 is negatively regulated in avian cells and acquires DNA-binding activity in certain cells upon heat shock.Heat shock genes are transcriptionally induced upon exposure of cells to physiological conditions, including stresses such as heat shock, heavy metals, oxidative stress, and amino acid analogs, and during nonstressful conditions such as cell growth, differentiation, and viral infections (27). The effects of heat shock and other stresses on heat shock gene transcription are mediated through interactions of the heat shock element (HSE) that is composed of at least three pentanucleotide modules (nGAAn) arranged as contiguous inverted repeats (33) with heat shock factor (HSF), the inducible transcriptional activator that regulates the transcription of these stress-inducible genes (25). In the budding yeasts Saccharomyces cerevisiae and Kluyveromyces lactis, HSF binds to HSE constitutively as a trimer and stimulates transcription upon heat shock (22,49,50). During deactivation of the heat shock response, the Cterminal activator domain is unmasked and becomes phosphorylated (9, 21, 32). In contrast, HSF does not bind to the HSE under normal growth conditions in the fission yeast Schizosaccharomyces pombe, Drosophila melanogaster, and higher eukaryotes but acquires HSE-binding activity by oligomerization to a trimer upon exposure to stress conditions (17,29,50,57,58,61,62). HSF trimers exhibit cooperative interactions between adjacent and distantly spaced HSEs as measured by DNA binding studies and transcriptional activation (3, 63).Recently, HSF genes from a number of eukaryotes have been cloned, thus allowing a comparative analysis to identify regulatory domains. The HSF family includes HSF1, HSF2, and HSF3 in higher eukaryotes ...

Impaired IgG Production in Mice Deficient for Heat Shock Transcription Factor 1

“…Whole cell extracts were prepared in buffer C (16). Aliquots containing 10 g of proteins were subjected to gel shift assay using an ideal HSE-oligonucleotide or an HSE2-oligonucleotide corresponding to the sequence of mouse IL-6 gene (Ϫ684 to Ϫ659) (20).…”

p53 Suppresses the c-Myb-induced Activation of Heat Shock Transcription Factor 3

“…The following binding buffers were used: 20 mM Hepes, pH 7.5, 125 mM NaCl, 1 mM EDTA, 0.1% Nonidet P-40, and 5 mM dithiothreitol for HSF3-p53 interaction; 20 mM Hepes, pH 7.5, 150 mM NaCl, 0.1% Nonidet P-40, and 1 mM dithiothreitol for interaction between Siah-1A and c-Myb. Co-immunoprecipitation-Rabbit anti-HSF3 antibody was described previously (18). To study the interaction among HSF3, c-Myb, and p53 (Figs.…”

Stress Protein HSP70 in Fish

“…The human, chicken, and tomato genomes contain multiple distinct hsf genes (Scharf et al 1990;Rabindran et al 1991;Sarge et al 1991;Schuetz et al 1991;Nakai and Morimoto 1993), whereas the genomes of yeast, Drosophila, and Xenopus each contain only a single hsf gene (Sorger and Pelham 1988;Wiederrecht et al 1988;Mercier et al 1997). In mammals and chickens, HSF1 and HSF3 were identified as rapidly activated, stress-responsive factors (Rabindran et al 1991;Sarge et al 1991;Nakai and Morimoto 1993;Nakai et al 1995). HSF2 responds to erythrocyte differentiation and preimplantation embryonic development (Hensold et al 1990;Schuetz et al 1991;Sistonen et al 1992), and HSF4 acts as both an activator and a repressor of tissue-specific heatshock genes through alternative splicing (Nakai et al 1997;Tanabe et al 1999).…”