Abstract
During heat shock (HS) and other stresses, HS gene transcription in eukaryotes is up-regulated by the transcription factor heat shock factor (HSF). While the identities of the major HS genes have been known for more than 30 years, it has been suspected that HSF binds to numerous other genes and potentially regulates their transcription. In this study, we have used a chromatin immunoprecipitation and microarray (ChIP-chip) approach to identify 434 regions in the Drosophila genome that are bound by HSF. We have also performed a transcript analysis of heat shocked Kc167 cells and third instar larvae and compared them to HSF binding sites. The heat-induced transcription profiles were quite different between cells and larvae and surprisingly only about 10% of the genes associated with HSF binding sites show changed transcription. There were also genes that showed changes in transcript levels that did not appear to correlate with HSF binding sites. Analysis of the locations of the HSF binding sites revealed that 57% were contained within genes with approximately 2/3rds of these sites being in introns. We also found that the insulator protein, BEAF, has enriched binding prior to HS to promoters of genes that are bound by HSF upon HS but that are not transcriptionally induced during HS. When the genes associated with HSF binding sites in promoters were analyzed for gene ontology terms, categories such as stress response and transferase activity were enriched whereas analysis of genes having HSF binding sites in introns identified those categories plus ones related to developmental processes and reproduction. These results suggest that Drosophila HSF may be regulating many genes besides the known HS genes and that some of these genes may be regulated during non-stress conditions.
Highlights
More than four decades ago Ritossa described a phenomenon where specific loci on the polytene chromosomes from third instar larvae of Drosophila decondensed or ‘‘puffed’’ when the larvae were exposed to heat or other forms of stress such as oxidative stress, inhibitors of respiration and certain metals [1]
We fixed both heat-shocked and non-shocked cells with formaldehyde to preserve protein-DNA interactions and immunoprecipitated Heat Shock Factor (HSF) bound chromatin complexes with an anti-HSF antibody generated by Westwood et al.,[32]
As a control for non-specific binding and for HSF binding under non-shock conditions, we performed a mock ChIP without antibody and an anti-HSF ChIP at room temperature ((RT), 22uC) respectively
Summary
More than four decades ago Ritossa described a phenomenon where specific loci on the polytene chromosomes from third instar larvae of Drosophila decondensed or ‘‘puffed’’ when the larvae were exposed to heat or other forms of stress such as oxidative stress, inhibitors of respiration and certain metals [1]. These puffs represented heat-induced sites of gene transcription and the genes residing there became known as the heat shock (HS) genes and their protein products the heat shock proteins (HSPs). In the metazoans studied far, binding of HSF or HSF1 to HSEs is low to virtually nonexistent in unshocked cells and upon HS or other stresses, HSF converts from a monomer to a trimeric form that binds to the HSEs with high affinity
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
