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STAT3 targets suggest mechanisms of aggressive tumorigenesis in diffuse large B-cell lymphoma.

Hardee J, Ouyang Z, Zhang Y, Kundaje A, Lacroute P, Snyder M - G3 (Bethesda) (2013)

Bottom Line: To investigate how STAT3 might contribute to this aggressive phenotype, we have integrated genome-wide studies of STAT3 DNA binding using chromatin immunoprecipitation-sequencing with whole-transcriptome profiling using RNA-sequencing.STAT3 binding sites are present near almost a third of all genes that differ in expression between the two subtypes, and examination of the affected genes identified previously undetected and clinically significant pathways downstream of STAT3 that drive oncogenesis.Novel treatments aimed at these pathways may increase the survivability of activated B-cell-like diffuse large B-cell lymphoma.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics, Stanford University School of Medicine, Stanford, California 94305.

ABSTRACT
The signal transducer and activator of transcription 3 (STAT3) is a transcription factor that, when dysregulated, becomes a powerful oncogene found in many human cancers, including diffuse large B-cell lymphoma. Diffuse large B-cell lymphoma is the most common form of non-Hodgkin's lymphoma and has two major subtypes: germinal center B-cell-like and activated B-cell-like. Compared with the germinal center B-cell-like form, activated B-cell-like lymphomas respond much more poorly to current therapies and often exhibit overexpression or overactivation of STAT3. To investigate how STAT3 might contribute to this aggressive phenotype, we have integrated genome-wide studies of STAT3 DNA binding using chromatin immunoprecipitation-sequencing with whole-transcriptome profiling using RNA-sequencing. STAT3 binding sites are present near almost a third of all genes that differ in expression between the two subtypes, and examination of the affected genes identified previously undetected and clinically significant pathways downstream of STAT3 that drive oncogenesis. Novel treatments aimed at these pathways may increase the survivability of activated B-cell-like diffuse large B-cell lymphoma.

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Hierarchical clustering analysis of STAT3 BRs. Clustering was performed on the 3524 regions that are differentially bound by STAT3 at FDR < 0.05 in ABC cell lines vs. GCB cell lines. Green and red blocks, respectively, represent high and low number of fragments sequenced in that BR relative to the average, whereas black blocks indicate no difference in expression. Fragment counts were log transformed to approximate a normal distribution, standardized by cell line, and each BR was normalized with sum of squares set to unity. GCB cell lines are represented by orange bars and ABC cell lines by blue bars. Generated using the heatmap.2 function in the R package gplots (R Development Core Team 2008; Warnes et al. 2011).
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fig2: Hierarchical clustering analysis of STAT3 BRs. Clustering was performed on the 3524 regions that are differentially bound by STAT3 at FDR < 0.05 in ABC cell lines vs. GCB cell lines. Green and red blocks, respectively, represent high and low number of fragments sequenced in that BR relative to the average, whereas black blocks indicate no difference in expression. Fragment counts were log transformed to approximate a normal distribution, standardized by cell line, and each BR was normalized with sum of squares set to unity. GCB cell lines are represented by orange bars and ABC cell lines by blue bars. Generated using the heatmap.2 function in the R package gplots (R Development Core Team 2008; Warnes et al. 2011).

Mentions: To identify target regions differentially bound by STAT3 in the ABC and GCB subtypes, we used the parameterized negative binomial model in DESeq (Anders and Huber 2010) to identify those BRs that displayed significantly different normalized fragment counts between the ABC and GCB cell lines, then corrected for multiple comparisons using the Benjamini-Hochberg procedure (Benjamini and Hochberg 1995). Of the 10,337 high-confidence BRs, one third (n = 3524) are differentially bound by STAT3 between the two subtypes at a false-discovery rate (FDR) < 0.05. (For a complete list of peaks by enriched subtype, see Table S4.) When these differentially bound peaks are clustered, the DLBCL cell lines cluster according to their GCB-ABC subtype (Figure 2). Consistently, more BRs are strongly bound in ABC than in GCB, although BRs with increased STAT3 binding occur surprisingly frequently in the GCB cell lines given their low level of STAT3 expression: 44% of differentially bound BRs (n = 1550) show more STAT3 binding in GCB, whereas 56% (n = 1974) are more strongly bound in ABC (Figure 3A). STAT3 protein levels are very low in GCB cell lines, so one possible explanation for its equivalent number of binding sites is simply that they represent lower occupancy regions. Indeed, we noticed the signal strength for the GCB lines was lower than that of the ABC cell lines and required additional ChIP-Seq replicate experiments to identify peaks.


STAT3 targets suggest mechanisms of aggressive tumorigenesis in diffuse large B-cell lymphoma.

Hardee J, Ouyang Z, Zhang Y, Kundaje A, Lacroute P, Snyder M - G3 (Bethesda) (2013)

Hierarchical clustering analysis of STAT3 BRs. Clustering was performed on the 3524 regions that are differentially bound by STAT3 at FDR < 0.05 in ABC cell lines vs. GCB cell lines. Green and red blocks, respectively, represent high and low number of fragments sequenced in that BR relative to the average, whereas black blocks indicate no difference in expression. Fragment counts were log transformed to approximate a normal distribution, standardized by cell line, and each BR was normalized with sum of squares set to unity. GCB cell lines are represented by orange bars and ABC cell lines by blue bars. Generated using the heatmap.2 function in the R package gplots (R Development Core Team 2008; Warnes et al. 2011).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3852380&req=5

fig2: Hierarchical clustering analysis of STAT3 BRs. Clustering was performed on the 3524 regions that are differentially bound by STAT3 at FDR < 0.05 in ABC cell lines vs. GCB cell lines. Green and red blocks, respectively, represent high and low number of fragments sequenced in that BR relative to the average, whereas black blocks indicate no difference in expression. Fragment counts were log transformed to approximate a normal distribution, standardized by cell line, and each BR was normalized with sum of squares set to unity. GCB cell lines are represented by orange bars and ABC cell lines by blue bars. Generated using the heatmap.2 function in the R package gplots (R Development Core Team 2008; Warnes et al. 2011).
Mentions: To identify target regions differentially bound by STAT3 in the ABC and GCB subtypes, we used the parameterized negative binomial model in DESeq (Anders and Huber 2010) to identify those BRs that displayed significantly different normalized fragment counts between the ABC and GCB cell lines, then corrected for multiple comparisons using the Benjamini-Hochberg procedure (Benjamini and Hochberg 1995). Of the 10,337 high-confidence BRs, one third (n = 3524) are differentially bound by STAT3 between the two subtypes at a false-discovery rate (FDR) < 0.05. (For a complete list of peaks by enriched subtype, see Table S4.) When these differentially bound peaks are clustered, the DLBCL cell lines cluster according to their GCB-ABC subtype (Figure 2). Consistently, more BRs are strongly bound in ABC than in GCB, although BRs with increased STAT3 binding occur surprisingly frequently in the GCB cell lines given their low level of STAT3 expression: 44% of differentially bound BRs (n = 1550) show more STAT3 binding in GCB, whereas 56% (n = 1974) are more strongly bound in ABC (Figure 3A). STAT3 protein levels are very low in GCB cell lines, so one possible explanation for its equivalent number of binding sites is simply that they represent lower occupancy regions. Indeed, we noticed the signal strength for the GCB lines was lower than that of the ABC cell lines and required additional ChIP-Seq replicate experiments to identify peaks.

Bottom Line: To investigate how STAT3 might contribute to this aggressive phenotype, we have integrated genome-wide studies of STAT3 DNA binding using chromatin immunoprecipitation-sequencing with whole-transcriptome profiling using RNA-sequencing.STAT3 binding sites are present near almost a third of all genes that differ in expression between the two subtypes, and examination of the affected genes identified previously undetected and clinically significant pathways downstream of STAT3 that drive oncogenesis.Novel treatments aimed at these pathways may increase the survivability of activated B-cell-like diffuse large B-cell lymphoma.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetics, Stanford University School of Medicine, Stanford, California 94305.

ABSTRACT
The signal transducer and activator of transcription 3 (STAT3) is a transcription factor that, when dysregulated, becomes a powerful oncogene found in many human cancers, including diffuse large B-cell lymphoma. Diffuse large B-cell lymphoma is the most common form of non-Hodgkin's lymphoma and has two major subtypes: germinal center B-cell-like and activated B-cell-like. Compared with the germinal center B-cell-like form, activated B-cell-like lymphomas respond much more poorly to current therapies and often exhibit overexpression or overactivation of STAT3. To investigate how STAT3 might contribute to this aggressive phenotype, we have integrated genome-wide studies of STAT3 DNA binding using chromatin immunoprecipitation-sequencing with whole-transcriptome profiling using RNA-sequencing. STAT3 binding sites are present near almost a third of all genes that differ in expression between the two subtypes, and examination of the affected genes identified previously undetected and clinically significant pathways downstream of STAT3 that drive oncogenesis. Novel treatments aimed at these pathways may increase the survivability of activated B-cell-like diffuse large B-cell lymphoma.

Show MeSH
Related in: MedlinePlus