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Regulatory network analyses reveal genome-wide potentiation of LIF signaling by glucocorticoids and define an innate cell defense response.

Langlais D, Couture C, Balsalobre A, Drouin J - PLoS Genet. (2008)

Bottom Line: Genome-wide ChIP-chip identified 3,449 STAT3 binding sites, whereas 2,396 genes regulated by LIF and/or Gc were found by expression profiling.Surprisingly, LIF on its own changed expression of only 85 genes but the joint action of LIF and Gc potentiated the expression of more than a thousand genes.Our analyses revealed an unexpected gene cluster that requires both stimuli for delayed activation; 83% of the genes in this cluster are involved in different cell defense mechanisms.

View Article: PubMed Central - PubMed

Affiliation: Laboratoire de Génétique Moléculaire, Institut de Recherches Cliniques de Montréal, Montréal, Quebec, Canada.

ABSTRACT
While the hypothalamo-pituitary-adrenal axis (HPA) activates a general stress response by increasing glucocorticoid (Gc) synthesis, biological stress resulting from infections triggers the inflammatory response through production of cytokines. The pituitary gland integrates some of these signals by responding to the pro-inflammatory cytokines IL6 and LIF and to a negative Gc feedback loop. The present work used whole-genome approaches to define the LIF/STAT3 regulatory network and to delineate cross-talk between this pathway and Gc action. Genome-wide ChIP-chip identified 3,449 STAT3 binding sites, whereas 2,396 genes regulated by LIF and/or Gc were found by expression profiling. Surprisingly, LIF on its own changed expression of only 85 genes but the joint action of LIF and Gc potentiated the expression of more than a thousand genes. Accordingly, activation of both LIF and Gc pathways also potentiated STAT3 and GR recruitment to many STAT3 targets. Our analyses revealed an unexpected gene cluster that requires both stimuli for delayed activation; 83% of the genes in this cluster are involved in different cell defense mechanisms. Thus, stressors that trigger both general stress and inflammatory responses lead to activation of a stereotypic innate cellular defense response.

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Related in: MedlinePlus

STAT3 genomic binding sites.A) Distances between STAT3 binding peaks determined by whole-genome ChIP-chip and nearest known genes (UCSC mm7 mouse genome assembly). Depending on position relative to the closest gene, data were computed as upstream to nearest TSS (left), relative to the TSS within the body of the gene itself (middle) or relative to the 3′ end of the gene (right). STAT3 binding sites that are outside these three categories were for the most part intergenic and this group constitutes 21.6% of the total number of STAT3 binding sites identified. B) Affymetrix Integrated Genome Browser (IGB) representation of tiling array data for STAT3 recruitment at the Pomc locus. In the top diagram, each vertical line represents the MAT score for one 25 bp oligonucleotide probe; each probe is spaced by 10 bp. The green solid horizontal bar indicates the interval of statistically significant (P≤10−5) STAT3 recruitment. This region contains a documented STAT3 binding site at −387/−379 bp [8],[9]. C) STAT3 binding sites within the Stat3/Stat5 locus. The upstream region of the Stat3 gene was previously shown to contain an auto-regulatory STAT3 binding site at position −338/−331 bp [26]. Strong recruitment of STAT3 was observed in this region but also at other positions within the Stat3/Stat5 locus. Statistically significant peaks (P≤10−5) of STAT3 binding are marked by the green boxes under the data diagram for tiling microarray data. D) Multiple STAT3 binding sites in the Socs3 locus including an upstream site that correlates with the previously documented site at −72/−62 bp [27]. E) STAT3 binding sites flanking a microRNA gene, miR-21. The STAT3 binding peak at −2801 bp is located near a STAT3 binding site previously identified in human [28]. F) WebLogo representation of known and computed preferred binding site for STAT3. The STAT3 binding site used by the MatBase database (Genomatix) for in silico analysis is shown together with a binding site derived from analysis of the 24 published sequences for STAT3 binding. All STAT3 binding regions from the tiling analysis were used to search for redundant DNA motifs, using non-biased algorithms: Consensus and AlignAce. As shown, both algorithms identified similar motifs. No other motif was identified within this dataset.
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pgen-1000224-g002: STAT3 genomic binding sites.A) Distances between STAT3 binding peaks determined by whole-genome ChIP-chip and nearest known genes (UCSC mm7 mouse genome assembly). Depending on position relative to the closest gene, data were computed as upstream to nearest TSS (left), relative to the TSS within the body of the gene itself (middle) or relative to the 3′ end of the gene (right). STAT3 binding sites that are outside these three categories were for the most part intergenic and this group constitutes 21.6% of the total number of STAT3 binding sites identified. B) Affymetrix Integrated Genome Browser (IGB) representation of tiling array data for STAT3 recruitment at the Pomc locus. In the top diagram, each vertical line represents the MAT score for one 25 bp oligonucleotide probe; each probe is spaced by 10 bp. The green solid horizontal bar indicates the interval of statistically significant (P≤10−5) STAT3 recruitment. This region contains a documented STAT3 binding site at −387/−379 bp [8],[9]. C) STAT3 binding sites within the Stat3/Stat5 locus. The upstream region of the Stat3 gene was previously shown to contain an auto-regulatory STAT3 binding site at position −338/−331 bp [26]. Strong recruitment of STAT3 was observed in this region but also at other positions within the Stat3/Stat5 locus. Statistically significant peaks (P≤10−5) of STAT3 binding are marked by the green boxes under the data diagram for tiling microarray data. D) Multiple STAT3 binding sites in the Socs3 locus including an upstream site that correlates with the previously documented site at −72/−62 bp [27]. E) STAT3 binding sites flanking a microRNA gene, miR-21. The STAT3 binding peak at −2801 bp is located near a STAT3 binding site previously identified in human [28]. F) WebLogo representation of known and computed preferred binding site for STAT3. The STAT3 binding site used by the MatBase database (Genomatix) for in silico analysis is shown together with a binding site derived from analysis of the 24 published sequences for STAT3 binding. All STAT3 binding regions from the tiling analysis were used to search for redundant DNA motifs, using non-biased algorithms: Consensus and AlignAce. As shown, both algorithms identified similar motifs. No other motif was identified within this dataset.

Mentions: The position of STAT3 binding sites on the mouse genome was analyzed relative to transcription start sites (TSS) of UCSC known genes. They were mapped either as upstream relative to known TSS, downstream from known TSS within the gene body or relative to the 3′ end of UCSC known genes (Figure 2A). This analysis clearly showed a preferential localization of STAT3 binding sites within 5 kb of TSS, with 19.4% of the total site number within this interval and 9.4% within 1 kb of TSS. Tiling array data for specific loci previously known to have STAT3 binding sites are also shown in Figure 2. For example, the promoter region of the Pomc gene is known to have a STAT3 binding site at −387/−379 bp [8]–[10], and the tiling array data show a peak of STAT3 recruitment over this promoter region (Figure 2B). Similarly, the promoter of the Stat3 gene itself is known to have a STAT3 binding site, and thus is subject to auto-regulation. The tiling array shows a peak of STAT3 recruitment (Figure 2C) that overlaps the reported STAT3 binding site at −338/−331 bp [26]. The Socs3 gene is involved in negative feedback regulation of STAT3 signaling and the Socs3 promoter has a STAT3 binding site at −64/−72 bp [27] that overlaps the observed peak of STAT3 recruitment (Figure 2D). In addition to these sites, the tiling array data revealed numerous other STAT3 binding sites in the Stat3/Stat5 and Socs3 loci; the biological relevance of these putative regulatory regions will need to be evaluated. Interestingly, STAT3 binding sites were found in close proximity to all Stat genes, except Stat6. Finally, STAT3 binding sites were found in the vicinity and promoter region of some microRNA genes, for example around the miR-21 gene (Figure 2E) that was implicated in the STAT3-dependent growth promotion activity of IL6 [28].


Regulatory network analyses reveal genome-wide potentiation of LIF signaling by glucocorticoids and define an innate cell defense response.

Langlais D, Couture C, Balsalobre A, Drouin J - PLoS Genet. (2008)

STAT3 genomic binding sites.A) Distances between STAT3 binding peaks determined by whole-genome ChIP-chip and nearest known genes (UCSC mm7 mouse genome assembly). Depending on position relative to the closest gene, data were computed as upstream to nearest TSS (left), relative to the TSS within the body of the gene itself (middle) or relative to the 3′ end of the gene (right). STAT3 binding sites that are outside these three categories were for the most part intergenic and this group constitutes 21.6% of the total number of STAT3 binding sites identified. B) Affymetrix Integrated Genome Browser (IGB) representation of tiling array data for STAT3 recruitment at the Pomc locus. In the top diagram, each vertical line represents the MAT score for one 25 bp oligonucleotide probe; each probe is spaced by 10 bp. The green solid horizontal bar indicates the interval of statistically significant (P≤10−5) STAT3 recruitment. This region contains a documented STAT3 binding site at −387/−379 bp [8],[9]. C) STAT3 binding sites within the Stat3/Stat5 locus. The upstream region of the Stat3 gene was previously shown to contain an auto-regulatory STAT3 binding site at position −338/−331 bp [26]. Strong recruitment of STAT3 was observed in this region but also at other positions within the Stat3/Stat5 locus. Statistically significant peaks (P≤10−5) of STAT3 binding are marked by the green boxes under the data diagram for tiling microarray data. D) Multiple STAT3 binding sites in the Socs3 locus including an upstream site that correlates with the previously documented site at −72/−62 bp [27]. E) STAT3 binding sites flanking a microRNA gene, miR-21. The STAT3 binding peak at −2801 bp is located near a STAT3 binding site previously identified in human [28]. F) WebLogo representation of known and computed preferred binding site for STAT3. The STAT3 binding site used by the MatBase database (Genomatix) for in silico analysis is shown together with a binding site derived from analysis of the 24 published sequences for STAT3 binding. All STAT3 binding regions from the tiling analysis were used to search for redundant DNA motifs, using non-biased algorithms: Consensus and AlignAce. As shown, both algorithms identified similar motifs. No other motif was identified within this dataset.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2562516&req=5

pgen-1000224-g002: STAT3 genomic binding sites.A) Distances between STAT3 binding peaks determined by whole-genome ChIP-chip and nearest known genes (UCSC mm7 mouse genome assembly). Depending on position relative to the closest gene, data were computed as upstream to nearest TSS (left), relative to the TSS within the body of the gene itself (middle) or relative to the 3′ end of the gene (right). STAT3 binding sites that are outside these three categories were for the most part intergenic and this group constitutes 21.6% of the total number of STAT3 binding sites identified. B) Affymetrix Integrated Genome Browser (IGB) representation of tiling array data for STAT3 recruitment at the Pomc locus. In the top diagram, each vertical line represents the MAT score for one 25 bp oligonucleotide probe; each probe is spaced by 10 bp. The green solid horizontal bar indicates the interval of statistically significant (P≤10−5) STAT3 recruitment. This region contains a documented STAT3 binding site at −387/−379 bp [8],[9]. C) STAT3 binding sites within the Stat3/Stat5 locus. The upstream region of the Stat3 gene was previously shown to contain an auto-regulatory STAT3 binding site at position −338/−331 bp [26]. Strong recruitment of STAT3 was observed in this region but also at other positions within the Stat3/Stat5 locus. Statistically significant peaks (P≤10−5) of STAT3 binding are marked by the green boxes under the data diagram for tiling microarray data. D) Multiple STAT3 binding sites in the Socs3 locus including an upstream site that correlates with the previously documented site at −72/−62 bp [27]. E) STAT3 binding sites flanking a microRNA gene, miR-21. The STAT3 binding peak at −2801 bp is located near a STAT3 binding site previously identified in human [28]. F) WebLogo representation of known and computed preferred binding site for STAT3. The STAT3 binding site used by the MatBase database (Genomatix) for in silico analysis is shown together with a binding site derived from analysis of the 24 published sequences for STAT3 binding. All STAT3 binding regions from the tiling analysis were used to search for redundant DNA motifs, using non-biased algorithms: Consensus and AlignAce. As shown, both algorithms identified similar motifs. No other motif was identified within this dataset.
Mentions: The position of STAT3 binding sites on the mouse genome was analyzed relative to transcription start sites (TSS) of UCSC known genes. They were mapped either as upstream relative to known TSS, downstream from known TSS within the gene body or relative to the 3′ end of UCSC known genes (Figure 2A). This analysis clearly showed a preferential localization of STAT3 binding sites within 5 kb of TSS, with 19.4% of the total site number within this interval and 9.4% within 1 kb of TSS. Tiling array data for specific loci previously known to have STAT3 binding sites are also shown in Figure 2. For example, the promoter region of the Pomc gene is known to have a STAT3 binding site at −387/−379 bp [8]–[10], and the tiling array data show a peak of STAT3 recruitment over this promoter region (Figure 2B). Similarly, the promoter of the Stat3 gene itself is known to have a STAT3 binding site, and thus is subject to auto-regulation. The tiling array shows a peak of STAT3 recruitment (Figure 2C) that overlaps the reported STAT3 binding site at −338/−331 bp [26]. The Socs3 gene is involved in negative feedback regulation of STAT3 signaling and the Socs3 promoter has a STAT3 binding site at −64/−72 bp [27] that overlaps the observed peak of STAT3 recruitment (Figure 2D). In addition to these sites, the tiling array data revealed numerous other STAT3 binding sites in the Stat3/Stat5 and Socs3 loci; the biological relevance of these putative regulatory regions will need to be evaluated. Interestingly, STAT3 binding sites were found in close proximity to all Stat genes, except Stat6. Finally, STAT3 binding sites were found in the vicinity and promoter region of some microRNA genes, for example around the miR-21 gene (Figure 2E) that was implicated in the STAT3-dependent growth promotion activity of IL6 [28].

Bottom Line: Genome-wide ChIP-chip identified 3,449 STAT3 binding sites, whereas 2,396 genes regulated by LIF and/or Gc were found by expression profiling.Surprisingly, LIF on its own changed expression of only 85 genes but the joint action of LIF and Gc potentiated the expression of more than a thousand genes.Our analyses revealed an unexpected gene cluster that requires both stimuli for delayed activation; 83% of the genes in this cluster are involved in different cell defense mechanisms.

View Article: PubMed Central - PubMed

Affiliation: Laboratoire de Génétique Moléculaire, Institut de Recherches Cliniques de Montréal, Montréal, Quebec, Canada.

ABSTRACT
While the hypothalamo-pituitary-adrenal axis (HPA) activates a general stress response by increasing glucocorticoid (Gc) synthesis, biological stress resulting from infections triggers the inflammatory response through production of cytokines. The pituitary gland integrates some of these signals by responding to the pro-inflammatory cytokines IL6 and LIF and to a negative Gc feedback loop. The present work used whole-genome approaches to define the LIF/STAT3 regulatory network and to delineate cross-talk between this pathway and Gc action. Genome-wide ChIP-chip identified 3,449 STAT3 binding sites, whereas 2,396 genes regulated by LIF and/or Gc were found by expression profiling. Surprisingly, LIF on its own changed expression of only 85 genes but the joint action of LIF and Gc potentiated the expression of more than a thousand genes. Accordingly, activation of both LIF and Gc pathways also potentiated STAT3 and GR recruitment to many STAT3 targets. Our analyses revealed an unexpected gene cluster that requires both stimuli for delayed activation; 83% of the genes in this cluster are involved in different cell defense mechanisms. Thus, stressors that trigger both general stress and inflammatory responses lead to activation of a stereotypic innate cellular defense response.

Show MeSH
Related in: MedlinePlus