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Genome-wide association of hypoxia-inducible factor (HIF)-1alpha and HIF-2alpha DNA binding with expression profiling of hypoxia-inducible transcripts.

Mole DR, Blancher C, Copley RR, Pollard PJ, Gleadle JM, Ragoussis J, Ratcliffe PJ - J. Biol. Chem. (2009)

Bottom Line: Comparison of HIF-promoter binding data with bidirectional HIF-dependent changes in transcript expression indicated that whereas a substantial proportion of positive responses (>20% across all significantly regulated genes) are direct, HIF-dependent gene suppression is almost entirely indirect.Comparison of HIF-1alpha- versus HIF-2alpha-binding sites revealed that whereas some loci bound HIF-1alpha in isolation, many bound both isoforms with similar affinity.Given emerging evidence for biologically distinct functions of HIF-1alpha versus HIF-2alpha understanding the mechanisms restricting HIF-2alpha activity will be of interest.

View Article: PubMed Central - PubMed

Affiliation: Henry Wellcome Building of Molecular Physiology, University of Oxford, Oxford OX3 7BN, United Kingdom. drmole@well.ox.ac.uk

ABSTRACT
Hypoxia-inducible factor (HIF) controls an extensive range of adaptive responses to hypoxia. To better understand this transcriptional cascade we performed genome-wide chromatin immunoprecipitation using antibodies to two major HIF-alpha subunits, and correlated the results with genome-wide transcript profiling. Within a tiled promoter array we identified 546 and 143 sequences that bound, respectively, to HIF-1alpha or HIF-2alpha at high stringency. Analysis of these sequences confirmed an identical core binding motif for HIF-1alpha and HIF-2alpha (RCGTG) but demonstrated that binding to this motif was highly selective, with binding enriched at distinct regions both upstream and downstream of the transcriptional start. Comparison of HIF-promoter binding data with bidirectional HIF-dependent changes in transcript expression indicated that whereas a substantial proportion of positive responses (>20% across all significantly regulated genes) are direct, HIF-dependent gene suppression is almost entirely indirect. Comparison of HIF-1alpha- versus HIF-2alpha-binding sites revealed that whereas some loci bound HIF-1alpha in isolation, many bound both isoforms with similar affinity. Despite high-affinity binding to multiple promoters, HIF-2alpha contributed to few, if any, of the transcriptional responses to acute hypoxia at these loci. Given emerging evidence for biologically distinct functions of HIF-1alpha versus HIF-2alpha understanding the mechanisms restricting HIF-2alpha activity will be of interest.

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Related in: MedlinePlus

Intersection between HIF-1α- and HIF-2α-binding gene loci. A, gene loci that bound either HIF-1α or HIF-2α or both isoforms with a Z-score >4. B, defining HIF-1α capture with a Z-score of 3 encompasses almost all HIF-2α binding loci. C, defining HIF-2α capture with a Z-score of 3 excludes almost one-third of HIF-1α binding loci.
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Figure 3: Intersection between HIF-1α- and HIF-2α-binding gene loci. A, gene loci that bound either HIF-1α or HIF-2α or both isoforms with a Z-score >4. B, defining HIF-1α capture with a Z-score of 3 encompasses almost all HIF-2α binding loci. C, defining HIF-2α capture with a Z-score of 3 excludes almost one-third of HIF-1α binding loci.

Mentions: Overall, there was a substantial overlap between the loci immunoprecipitated with the two HIF isoforms. Thus, of the 394 loci that bound HIF-1α and the 134 loci that bound HIF-2α, 90 bound both isoforms. When the stringency of binding for the second isoform was relaxed slightly (MA score >3), the overlap increased, with 250/394 HIF-1α loci binding HIF-2α and 130/134 HIF-2α loci binding HIF-1α. In addition, when both HIF isoforms bind to the same gene locus, they do so at a common site. Thus, of the 90 gene loci that bound both HIF isoforms at the highest level of stringency, all apart from 3 bound HIF-1α and HIF-2α at overlapping sites (Fig. 3).


Genome-wide association of hypoxia-inducible factor (HIF)-1alpha and HIF-2alpha DNA binding with expression profiling of hypoxia-inducible transcripts.

Mole DR, Blancher C, Copley RR, Pollard PJ, Gleadle JM, Ragoussis J, Ratcliffe PJ - J. Biol. Chem. (2009)

Intersection between HIF-1α- and HIF-2α-binding gene loci. A, gene loci that bound either HIF-1α or HIF-2α or both isoforms with a Z-score >4. B, defining HIF-1α capture with a Z-score of 3 encompasses almost all HIF-2α binding loci. C, defining HIF-2α capture with a Z-score of 3 excludes almost one-third of HIF-1α binding loci.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2719312&req=5

Figure 3: Intersection between HIF-1α- and HIF-2α-binding gene loci. A, gene loci that bound either HIF-1α or HIF-2α or both isoforms with a Z-score >4. B, defining HIF-1α capture with a Z-score of 3 encompasses almost all HIF-2α binding loci. C, defining HIF-2α capture with a Z-score of 3 excludes almost one-third of HIF-1α binding loci.
Mentions: Overall, there was a substantial overlap between the loci immunoprecipitated with the two HIF isoforms. Thus, of the 394 loci that bound HIF-1α and the 134 loci that bound HIF-2α, 90 bound both isoforms. When the stringency of binding for the second isoform was relaxed slightly (MA score >3), the overlap increased, with 250/394 HIF-1α loci binding HIF-2α and 130/134 HIF-2α loci binding HIF-1α. In addition, when both HIF isoforms bind to the same gene locus, they do so at a common site. Thus, of the 90 gene loci that bound both HIF isoforms at the highest level of stringency, all apart from 3 bound HIF-1α and HIF-2α at overlapping sites (Fig. 3).

Bottom Line: Comparison of HIF-promoter binding data with bidirectional HIF-dependent changes in transcript expression indicated that whereas a substantial proportion of positive responses (>20% across all significantly regulated genes) are direct, HIF-dependent gene suppression is almost entirely indirect.Comparison of HIF-1alpha- versus HIF-2alpha-binding sites revealed that whereas some loci bound HIF-1alpha in isolation, many bound both isoforms with similar affinity.Given emerging evidence for biologically distinct functions of HIF-1alpha versus HIF-2alpha understanding the mechanisms restricting HIF-2alpha activity will be of interest.

View Article: PubMed Central - PubMed

Affiliation: Henry Wellcome Building of Molecular Physiology, University of Oxford, Oxford OX3 7BN, United Kingdom. drmole@well.ox.ac.uk

ABSTRACT
Hypoxia-inducible factor (HIF) controls an extensive range of adaptive responses to hypoxia. To better understand this transcriptional cascade we performed genome-wide chromatin immunoprecipitation using antibodies to two major HIF-alpha subunits, and correlated the results with genome-wide transcript profiling. Within a tiled promoter array we identified 546 and 143 sequences that bound, respectively, to HIF-1alpha or HIF-2alpha at high stringency. Analysis of these sequences confirmed an identical core binding motif for HIF-1alpha and HIF-2alpha (RCGTG) but demonstrated that binding to this motif was highly selective, with binding enriched at distinct regions both upstream and downstream of the transcriptional start. Comparison of HIF-promoter binding data with bidirectional HIF-dependent changes in transcript expression indicated that whereas a substantial proportion of positive responses (>20% across all significantly regulated genes) are direct, HIF-dependent gene suppression is almost entirely indirect. Comparison of HIF-1alpha- versus HIF-2alpha-binding sites revealed that whereas some loci bound HIF-1alpha in isolation, many bound both isoforms with similar affinity. Despite high-affinity binding to multiple promoters, HIF-2alpha contributed to few, if any, of the transcriptional responses to acute hypoxia at these loci. Given emerging evidence for biologically distinct functions of HIF-1alpha versus HIF-2alpha understanding the mechanisms restricting HIF-2alpha activity will be of interest.

Show MeSH
Related in: MedlinePlus