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Whole-genome analysis reveals that active heat shock factor binding sites are mostly associated with non-heat shock genes in Drosophila melanogaster.

Gonsalves SE, Moses AM, Razak Z, Robert F, Westwood JT - PLoS ONE (2011)

Bottom Line: There were also genes that showed changes in transcript levels that did not appear to correlate with HSF binding sites.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.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.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell and Systems Biology, University of Toronto, Mississauga, Canada.

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.

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HSF binds to introns in the three major ecdysone-inducible genes, Eip75B, Eip74EF, and br.We have identified six HSF-binding sites within the bodies of these three genes. Each site is located within an intron and overlaps with a binding sites for the ecdysone receptor complex (EcR-C) [51]. Five out the six sites are also located within 5 kb of the transcription start site of one or more isoforms and the sixth site is still only 10 kb away from a transcription start site. Each orange bar represents a probe from the tiling array that is part of an HSF bound segment and its height indicates its fold enrichment relative to WCE. As with Figure 4, this image was generated using the UCSC Genome Browser to illustrate the chromosome region represented in bp (according to release 4.2 of the Drosophila genome) as indicated at the top and the legend is the same as that in Figure 4.
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pone-0015934-g011: HSF binds to introns in the three major ecdysone-inducible genes, Eip75B, Eip74EF, and br.We have identified six HSF-binding sites within the bodies of these three genes. Each site is located within an intron and overlaps with a binding sites for the ecdysone receptor complex (EcR-C) [51]. Five out the six sites are also located within 5 kb of the transcription start site of one or more isoforms and the sixth site is still only 10 kb away from a transcription start site. Each orange bar represents a probe from the tiling array that is part of an HSF bound segment and its height indicates its fold enrichment relative to WCE. As with Figure 4, this image was generated using the UCSC Genome Browser to illustrate the chromosome region represented in bp (according to release 4.2 of the Drosophila genome) as indicated at the top and the legend is the same as that in Figure 4.

Mentions: Since HSF binds a large number of introns of genes that do not appear to be transcriptionally induced during HS, we decided to examine a subset of HSF-bound introns more closely. The functional enrichment analysis presented in the previous section (Figure 8), indicates that intron-bound genes are strongly enriched in functions related to both the regulation of transcription and to developmental processes. We had noted in a previous study that HSF binds to two well examined chromosomal loci, 74EF and 75B, the sites of two of the early ecdysone inducible transcription factor genes, Eip74EF and Eip75B [32]. These two genes together with br, are major components of the transcription hierarchy that are involved in the developmental response to ecdysone by regulating the transcription of a large set of secondary ecdysone-response genes (for review see [50]). All three of these genes are very large in size spanning from 59 to over 100 kb and each encodes multiple isoforms with at least two distinct transcription start sites (for review see [50]). Within these three genes, we have found a total of six HSF-binding sites, four within Eip75B, one within Eip74EF and one within br (Figure 11). Interestingly, all but one of these intronic HSF-binding sites are located less than 5 kb upstream of 5′ end of one or more isoforms. Given that introns in these genes that are bound by HSF are rather large (for example the first intron of Eip75B in which HSF occupies two sites, is over 60 kb in length and the intron bound by HSF in Eip74EF is over 20 kb) this distribution of HSF-binding sites appears to exhibit a strong bias for the extreme 3′ end of the introns close to the 5′ ends of alternate isoforms. In addition, all five HSF binding sites overlap with binding sites identified in Kc167 cells for the ecdysone receptor complex (EcR-C) [51], the nuclear receptor complex responsible for the stage and tissue specific activation of Eip75B, Eip74EF, and br [52], [53].


Whole-genome analysis reveals that active heat shock factor binding sites are mostly associated with non-heat shock genes in Drosophila melanogaster.

Gonsalves SE, Moses AM, Razak Z, Robert F, Westwood JT - PLoS ONE (2011)

HSF binds to introns in the three major ecdysone-inducible genes, Eip75B, Eip74EF, and br.We have identified six HSF-binding sites within the bodies of these three genes. Each site is located within an intron and overlaps with a binding sites for the ecdysone receptor complex (EcR-C) [51]. Five out the six sites are also located within 5 kb of the transcription start site of one or more isoforms and the sixth site is still only 10 kb away from a transcription start site. Each orange bar represents a probe from the tiling array that is part of an HSF bound segment and its height indicates its fold enrichment relative to WCE. As with Figure 4, this image was generated using the UCSC Genome Browser to illustrate the chromosome region represented in bp (according to release 4.2 of the Drosophila genome) as indicated at the top and the legend is the same as that in Figure 4.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3021535&req=5

pone-0015934-g011: HSF binds to introns in the three major ecdysone-inducible genes, Eip75B, Eip74EF, and br.We have identified six HSF-binding sites within the bodies of these three genes. Each site is located within an intron and overlaps with a binding sites for the ecdysone receptor complex (EcR-C) [51]. Five out the six sites are also located within 5 kb of the transcription start site of one or more isoforms and the sixth site is still only 10 kb away from a transcription start site. Each orange bar represents a probe from the tiling array that is part of an HSF bound segment and its height indicates its fold enrichment relative to WCE. As with Figure 4, this image was generated using the UCSC Genome Browser to illustrate the chromosome region represented in bp (according to release 4.2 of the Drosophila genome) as indicated at the top and the legend is the same as that in Figure 4.
Mentions: Since HSF binds a large number of introns of genes that do not appear to be transcriptionally induced during HS, we decided to examine a subset of HSF-bound introns more closely. The functional enrichment analysis presented in the previous section (Figure 8), indicates that intron-bound genes are strongly enriched in functions related to both the regulation of transcription and to developmental processes. We had noted in a previous study that HSF binds to two well examined chromosomal loci, 74EF and 75B, the sites of two of the early ecdysone inducible transcription factor genes, Eip74EF and Eip75B [32]. These two genes together with br, are major components of the transcription hierarchy that are involved in the developmental response to ecdysone by regulating the transcription of a large set of secondary ecdysone-response genes (for review see [50]). All three of these genes are very large in size spanning from 59 to over 100 kb and each encodes multiple isoforms with at least two distinct transcription start sites (for review see [50]). Within these three genes, we have found a total of six HSF-binding sites, four within Eip75B, one within Eip74EF and one within br (Figure 11). Interestingly, all but one of these intronic HSF-binding sites are located less than 5 kb upstream of 5′ end of one or more isoforms. Given that introns in these genes that are bound by HSF are rather large (for example the first intron of Eip75B in which HSF occupies two sites, is over 60 kb in length and the intron bound by HSF in Eip74EF is over 20 kb) this distribution of HSF-binding sites appears to exhibit a strong bias for the extreme 3′ end of the introns close to the 5′ ends of alternate isoforms. In addition, all five HSF binding sites overlap with binding sites identified in Kc167 cells for the ecdysone receptor complex (EcR-C) [51], the nuclear receptor complex responsible for the stage and tissue specific activation of Eip75B, Eip74EF, and br [52], [53].

Bottom Line: There were also genes that showed changes in transcript levels that did not appear to correlate with HSF binding sites.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.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.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell and Systems Biology, University of Toronto, Mississauga, Canada.

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.

Show MeSH
Related in: MedlinePlus