Limits...
A large gene network in immature erythroid cells is controlled by the myeloid and B cell transcriptional regulator PU.1.

Wontakal SN, Guo X, Will B, Shi M, Raha D, Mahajan MC, Weissman S, Snyder M, Steidl U, Zheng D, Skoultchi AI - PLoS Genet. (2011)

Bottom Line: PU.1 is also expressed in erythroid progenitors, where it blocks erythroid differentiation by binding to and inhibiting the main erythroid promoting factor, GATA-1.By analyzing fetal liver erythroid progenitors from mice with low PU.1 expression, we also show that the earliest erythroid committed cells are dramatically reduced in vivo.Furthermore, we find that PU.1 also regulates many of the same genes and pathways in other blood cells, leading us to propose that PU.1 is a multifaceted factor with overlapping, as well as distinct, functions in several hematopoietic lineages.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, Albert Einstein College of Medicine, Bronx, New York, United States of America.

ABSTRACT
PU.1 is a hematopoietic transcription factor that is required for the development of myeloid and B cells. PU.1 is also expressed in erythroid progenitors, where it blocks erythroid differentiation by binding to and inhibiting the main erythroid promoting factor, GATA-1. However, other mechanisms by which PU.1 affects the fate of erythroid progenitors have not been thoroughly explored. Here, we used ChIP-Seq analysis for PU.1 and gene expression profiling in erythroid cells to show that PU.1 regulates an extensive network of genes that constitute major pathways for controlling growth and survival of immature erythroid cells. By analyzing fetal liver erythroid progenitors from mice with low PU.1 expression, we also show that the earliest erythroid committed cells are dramatically reduced in vivo. Furthermore, we find that PU.1 also regulates many of the same genes and pathways in other blood cells, leading us to propose that PU.1 is a multifaceted factor with overlapping, as well as distinct, functions in several hematopoietic lineages.

Show MeSH
Properties of the PU.1 ChIP-Seq peaks in ES-EP and MEL cells.(A) Sample signal tracks of PU.1 ChIP-Seq data from MEL cells and ES-EP are shown for an ∼500 kb region of chromosome 15 in the Integrated Genome Browser (IGB) (Affymetrix), with the y-axis representing the number of reads. Input DNA controls are also shown for both cell types. (B) Sequence logos for the enriched motifs within PU.1 ChIP-Seq peaks from MEL (left) and ES-EP (right) cells, derived from MEME motif analysis (see Materials and Methods). (C) The distance between each PU.1 ChIP-Seq peak and the TSS within 2 kb was computed and the results are binned and plotted for MEL (top left) and ES-EP (bottom left). Peaks were further annotated by their genomic locations with respect to current gene annotation (right) and classified as proximal promoter (+/−2 kb of TSS), 3′ end of gene (+/−2 kb of TES), gene body (between +2 kb of TSS and −2 kb of TES), miRNA promoters, or otherwise intergenic regions.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3111485&req=5

pgen-1001392-g002: Properties of the PU.1 ChIP-Seq peaks in ES-EP and MEL cells.(A) Sample signal tracks of PU.1 ChIP-Seq data from MEL cells and ES-EP are shown for an ∼500 kb region of chromosome 15 in the Integrated Genome Browser (IGB) (Affymetrix), with the y-axis representing the number of reads. Input DNA controls are also shown for both cell types. (B) Sequence logos for the enriched motifs within PU.1 ChIP-Seq peaks from MEL (left) and ES-EP (right) cells, derived from MEME motif analysis (see Materials and Methods). (C) The distance between each PU.1 ChIP-Seq peak and the TSS within 2 kb was computed and the results are binned and plotted for MEL (top left) and ES-EP (bottom left). Peaks were further annotated by their genomic locations with respect to current gene annotation (right) and classified as proximal promoter (+/−2 kb of TSS), 3′ end of gene (+/−2 kb of TES), gene body (between +2 kb of TSS and −2 kb of TES), miRNA promoters, or otherwise intergenic regions.

Mentions: As a first step in identifying the transcriptional network controlled by PU.1 in immature erythroid cells, we performed ChIP-Seq in normal proliferating erythroid progenitors derived from embryonic stem cells (ES-EP) [23] and leukemic erythroblasts (MEL cells). We obtained a total of 13,416,531 and 12,710,420 uniquely mapped reads in ES-EP and MEL cells, respectively. Using two peak calling programs, cisGenome and spp [24], [25], we identified a total of 16,241 peaks of PU.1 occupancy in ES-EP and 16,599 peaks in MEL cells. We also compared the number of reads in a given peak in MEL cells and ES-EP and then assigned a peak as present in both cell types (≤5 fold difference in the number of reads) or enriched in one cell type (>5 fold difference) (Figure 1A). With this classification scheme, we identified 16,011 peaks that are shared between the two cell types and 230 and 588 peaks that are enriched in ES-EP and MEL cells, respectively (Figure 1B left). Strikingly, more than 95% of the peaks are shared between the two cell types (Figure 1B left). Statistical analysis of the data presented in Figure 1A also revealed a strong similarity between PU.1 binding in the two cell types as evidenced by a correlation coefficient of 0.800 (p-value<2.2×10−16). This similarity is even visually evident from examination of the signal tracks of 500 kb windows, such as the one displayed in Figure 2A. We used qChIP to verify that some of the rare loci enriched in one cell type are indeed differentially occupied in the two cell types (Figure S1). Overall, our results indicate that the binding patterns of PU.1 are highly similar in normal and leukemic erythroid cells.


A large gene network in immature erythroid cells is controlled by the myeloid and B cell transcriptional regulator PU.1.

Wontakal SN, Guo X, Will B, Shi M, Raha D, Mahajan MC, Weissman S, Snyder M, Steidl U, Zheng D, Skoultchi AI - PLoS Genet. (2011)

Properties of the PU.1 ChIP-Seq peaks in ES-EP and MEL cells.(A) Sample signal tracks of PU.1 ChIP-Seq data from MEL cells and ES-EP are shown for an ∼500 kb region of chromosome 15 in the Integrated Genome Browser (IGB) (Affymetrix), with the y-axis representing the number of reads. Input DNA controls are also shown for both cell types. (B) Sequence logos for the enriched motifs within PU.1 ChIP-Seq peaks from MEL (left) and ES-EP (right) cells, derived from MEME motif analysis (see Materials and Methods). (C) The distance between each PU.1 ChIP-Seq peak and the TSS within 2 kb was computed and the results are binned and plotted for MEL (top left) and ES-EP (bottom left). Peaks were further annotated by their genomic locations with respect to current gene annotation (right) and classified as proximal promoter (+/−2 kb of TSS), 3′ end of gene (+/−2 kb of TES), gene body (between +2 kb of TSS and −2 kb of TES), miRNA promoters, or otherwise intergenic regions.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3111485&req=5

pgen-1001392-g002: Properties of the PU.1 ChIP-Seq peaks in ES-EP and MEL cells.(A) Sample signal tracks of PU.1 ChIP-Seq data from MEL cells and ES-EP are shown for an ∼500 kb region of chromosome 15 in the Integrated Genome Browser (IGB) (Affymetrix), with the y-axis representing the number of reads. Input DNA controls are also shown for both cell types. (B) Sequence logos for the enriched motifs within PU.1 ChIP-Seq peaks from MEL (left) and ES-EP (right) cells, derived from MEME motif analysis (see Materials and Methods). (C) The distance between each PU.1 ChIP-Seq peak and the TSS within 2 kb was computed and the results are binned and plotted for MEL (top left) and ES-EP (bottom left). Peaks were further annotated by their genomic locations with respect to current gene annotation (right) and classified as proximal promoter (+/−2 kb of TSS), 3′ end of gene (+/−2 kb of TES), gene body (between +2 kb of TSS and −2 kb of TES), miRNA promoters, or otherwise intergenic regions.
Mentions: As a first step in identifying the transcriptional network controlled by PU.1 in immature erythroid cells, we performed ChIP-Seq in normal proliferating erythroid progenitors derived from embryonic stem cells (ES-EP) [23] and leukemic erythroblasts (MEL cells). We obtained a total of 13,416,531 and 12,710,420 uniquely mapped reads in ES-EP and MEL cells, respectively. Using two peak calling programs, cisGenome and spp [24], [25], we identified a total of 16,241 peaks of PU.1 occupancy in ES-EP and 16,599 peaks in MEL cells. We also compared the number of reads in a given peak in MEL cells and ES-EP and then assigned a peak as present in both cell types (≤5 fold difference in the number of reads) or enriched in one cell type (>5 fold difference) (Figure 1A). With this classification scheme, we identified 16,011 peaks that are shared between the two cell types and 230 and 588 peaks that are enriched in ES-EP and MEL cells, respectively (Figure 1B left). Strikingly, more than 95% of the peaks are shared between the two cell types (Figure 1B left). Statistical analysis of the data presented in Figure 1A also revealed a strong similarity between PU.1 binding in the two cell types as evidenced by a correlation coefficient of 0.800 (p-value<2.2×10−16). This similarity is even visually evident from examination of the signal tracks of 500 kb windows, such as the one displayed in Figure 2A. We used qChIP to verify that some of the rare loci enriched in one cell type are indeed differentially occupied in the two cell types (Figure S1). Overall, our results indicate that the binding patterns of PU.1 are highly similar in normal and leukemic erythroid cells.

Bottom Line: PU.1 is also expressed in erythroid progenitors, where it blocks erythroid differentiation by binding to and inhibiting the main erythroid promoting factor, GATA-1.By analyzing fetal liver erythroid progenitors from mice with low PU.1 expression, we also show that the earliest erythroid committed cells are dramatically reduced in vivo.Furthermore, we find that PU.1 also regulates many of the same genes and pathways in other blood cells, leading us to propose that PU.1 is a multifaceted factor with overlapping, as well as distinct, functions in several hematopoietic lineages.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, Albert Einstein College of Medicine, Bronx, New York, United States of America.

ABSTRACT
PU.1 is a hematopoietic transcription factor that is required for the development of myeloid and B cells. PU.1 is also expressed in erythroid progenitors, where it blocks erythroid differentiation by binding to and inhibiting the main erythroid promoting factor, GATA-1. However, other mechanisms by which PU.1 affects the fate of erythroid progenitors have not been thoroughly explored. Here, we used ChIP-Seq analysis for PU.1 and gene expression profiling in erythroid cells to show that PU.1 regulates an extensive network of genes that constitute major pathways for controlling growth and survival of immature erythroid cells. By analyzing fetal liver erythroid progenitors from mice with low PU.1 expression, we also show that the earliest erythroid committed cells are dramatically reduced in vivo. Furthermore, we find that PU.1 also regulates many of the same genes and pathways in other blood cells, leading us to propose that PU.1 is a multifaceted factor with overlapping, as well as distinct, functions in several hematopoietic lineages.

Show MeSH