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Cis-regulatory mechanisms governing stem and progenitor cell transitions.

Johnson KD, Kong G, Gao X, Chang YI, Hewitt KJ, Sanalkumar R, Prathibha R, Ranheim EA, Dewey CN, Zhang J, Bresnick EH - Sci Adv (2015)

Bottom Line: The -77 established a vital sector of the myeloid progenitor transcriptome, conferring multipotentiality.Unlike the +9.5(-/-) embryos, hematopoietic stem cell genesis was unaffected in -77(-/-) embryos.These results illustrate a paradigm in which cis-elements in a locus differentially control stem and progenitor cell transitions, and therefore the individual cis-element alterations cause unique and overlapping disease phenotypes.

View Article: PubMed Central - PubMed

Affiliation: Carbone Cancer Center, Department of Cell and Regenerative Biology, University of Wisconsin School of Medicine and Public Health, Madison, WI 53705, USA. ; University of Wisconsin-Madison Blood Research Program, Madison, WI 53705, USA.

ABSTRACT
Cis-element encyclopedias provide information on phenotypic diversity and disease mechanisms. Although cis-element polymorphisms and mutations are instructive, deciphering function remains challenging. Mutation of an intronic GATA motif (+9.5) in GATA2, encoding a master regulator of hematopoiesis, underlies an immunodeficiency associated with myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML). Whereas an inversion relocalizes another GATA2 cis-element (-77) to the proto-oncogene EVI1, inducing EVI1 expression and AML, whether this reflects ectopic or physiological activity is unknown. We describe a mouse strain that decouples -77 function from proto-oncogene deregulation. The -77(-/-) mice exhibited a novel phenotypic constellation including late embryonic lethality and anemia. The -77 established a vital sector of the myeloid progenitor transcriptome, conferring multipotentiality. Unlike the +9.5(-/-) embryos, hematopoietic stem cell genesis was unaffected in -77(-/-) embryos. These results illustrate a paradigm in which cis-elements in a locus differentially control stem and progenitor cell transitions, and therefore the individual cis-element alterations cause unique and overlapping disease phenotypes.

No MeSH data available.


Related in: MedlinePlus

−77 establishes a functionally vital sector of the myeloid progenitor cell transcriptome.(A) Quantitative gene expression analysis in CMPs, GMPs, and MEPs from −77+/+ and 77−/− E13.5 fetal livers [−77+/+ (n = 4) and −77−/− (n = 4)]. (B) MA plot of RNA-seq–based comparison of CMP transcriptomes from −77+/+ and −77−/− E13.5 fetal livers [−77+/+ (n = 3) and −77−/− (n = 3)]. Red points indicate down- or up-regulated genes [false discovery rate (FDR) <0.05]. (C) Heatmap depicting statistically significant genes down- or up-regulated by >1.5-fold. (D) Venn diagram depicting the extent of overlap between genes regulated by +9.5 in AGM (13) and −77 in CMPs. Genes shared between data sets are indicated in red in (C). (E) Evidence for direct GATA-2 regulation of −77 target genes. ChIP-seq profiles of GATA-2 and Scl/TAL1 in HPC-7 cells (58) and quantitative gene expression analysis in CMPs from −77+/+ and −77−/− E13.5 fetal livers [−77+/+ (n = 4) and −77−/− (n = 4)]. Graphs show means ± SEM; *P < 0.05, **P < 0.01, ***P < 0.001.
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Figure 5: −77 establishes a functionally vital sector of the myeloid progenitor cell transcriptome.(A) Quantitative gene expression analysis in CMPs, GMPs, and MEPs from −77+/+ and 77−/− E13.5 fetal livers [−77+/+ (n = 4) and −77−/− (n = 4)]. (B) MA plot of RNA-seq–based comparison of CMP transcriptomes from −77+/+ and −77−/− E13.5 fetal livers [−77+/+ (n = 3) and −77−/− (n = 3)]. Red points indicate down- or up-regulated genes [false discovery rate (FDR) <0.05]. (C) Heatmap depicting statistically significant genes down- or up-regulated by >1.5-fold. (D) Venn diagram depicting the extent of overlap between genes regulated by +9.5 in AGM (13) and −77 in CMPs. Genes shared between data sets are indicated in red in (C). (E) Evidence for direct GATA-2 regulation of −77 target genes. ChIP-seq profiles of GATA-2 and Scl/TAL1 in HPC-7 cells (58) and quantitative gene expression analysis in CMPs from −77+/+ and −77−/− E13.5 fetal livers [−77+/+ (n = 4) and −77−/− (n = 4)]. Graphs show means ± SEM; *P < 0.05, **P < 0.01, ***P < 0.001.

Mentions: As the −77 mutation reduced myeloid progenitor abundance and impaired function, we tested whether the mutation altered the expression of established regulators of myeloerythroid differentiation. Consistent with the MEP loss, Gata1 expression was reduced in CMPs and GMPs, and Zfpm1 (FOG-1) expression was reduced in CMPs (Fig. 5A). In the very few −77−/− MEPs produced, the expression of both genes was unaffected. Among myeloid regulatory factors, −77−/− progenitors expressed normal levels of Sfpi1 (PU.1) and C/EBPα (CCAAT/enhancer binding protein-α), whereas growth factor independence 1 (Gfi1) was reduced 2.6-fold selectively in GMPs. The expression of pre–B cell leukemia homeobox 1 (Pbx1), a proto-oncogene that regulates CMP levels and differentiation potential in the fetal liver (33), was reduced three-fold in CMPs but was increased significantly in GMPs. Although the CMP compartment was substantially reduced in Pbx1−/− embryos but increased in −77−/− fetal livers, both mutations similarly altered the CMP colony-forming potential, leading to reduced erythroid colonies and increased monocytes.


Cis-regulatory mechanisms governing stem and progenitor cell transitions.

Johnson KD, Kong G, Gao X, Chang YI, Hewitt KJ, Sanalkumar R, Prathibha R, Ranheim EA, Dewey CN, Zhang J, Bresnick EH - Sci Adv (2015)

−77 establishes a functionally vital sector of the myeloid progenitor cell transcriptome.(A) Quantitative gene expression analysis in CMPs, GMPs, and MEPs from −77+/+ and 77−/− E13.5 fetal livers [−77+/+ (n = 4) and −77−/− (n = 4)]. (B) MA plot of RNA-seq–based comparison of CMP transcriptomes from −77+/+ and −77−/− E13.5 fetal livers [−77+/+ (n = 3) and −77−/− (n = 3)]. Red points indicate down- or up-regulated genes [false discovery rate (FDR) <0.05]. (C) Heatmap depicting statistically significant genes down- or up-regulated by >1.5-fold. (D) Venn diagram depicting the extent of overlap between genes regulated by +9.5 in AGM (13) and −77 in CMPs. Genes shared between data sets are indicated in red in (C). (E) Evidence for direct GATA-2 regulation of −77 target genes. ChIP-seq profiles of GATA-2 and Scl/TAL1 in HPC-7 cells (58) and quantitative gene expression analysis in CMPs from −77+/+ and −77−/− E13.5 fetal livers [−77+/+ (n = 4) and −77−/− (n = 4)]. Graphs show means ± SEM; *P < 0.05, **P < 0.01, ***P < 0.001.
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Related In: Results  -  Collection

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Figure 5: −77 establishes a functionally vital sector of the myeloid progenitor cell transcriptome.(A) Quantitative gene expression analysis in CMPs, GMPs, and MEPs from −77+/+ and 77−/− E13.5 fetal livers [−77+/+ (n = 4) and −77−/− (n = 4)]. (B) MA plot of RNA-seq–based comparison of CMP transcriptomes from −77+/+ and −77−/− E13.5 fetal livers [−77+/+ (n = 3) and −77−/− (n = 3)]. Red points indicate down- or up-regulated genes [false discovery rate (FDR) <0.05]. (C) Heatmap depicting statistically significant genes down- or up-regulated by >1.5-fold. (D) Venn diagram depicting the extent of overlap between genes regulated by +9.5 in AGM (13) and −77 in CMPs. Genes shared between data sets are indicated in red in (C). (E) Evidence for direct GATA-2 regulation of −77 target genes. ChIP-seq profiles of GATA-2 and Scl/TAL1 in HPC-7 cells (58) and quantitative gene expression analysis in CMPs from −77+/+ and −77−/− E13.5 fetal livers [−77+/+ (n = 4) and −77−/− (n = 4)]. Graphs show means ± SEM; *P < 0.05, **P < 0.01, ***P < 0.001.
Mentions: As the −77 mutation reduced myeloid progenitor abundance and impaired function, we tested whether the mutation altered the expression of established regulators of myeloerythroid differentiation. Consistent with the MEP loss, Gata1 expression was reduced in CMPs and GMPs, and Zfpm1 (FOG-1) expression was reduced in CMPs (Fig. 5A). In the very few −77−/− MEPs produced, the expression of both genes was unaffected. Among myeloid regulatory factors, −77−/− progenitors expressed normal levels of Sfpi1 (PU.1) and C/EBPα (CCAAT/enhancer binding protein-α), whereas growth factor independence 1 (Gfi1) was reduced 2.6-fold selectively in GMPs. The expression of pre–B cell leukemia homeobox 1 (Pbx1), a proto-oncogene that regulates CMP levels and differentiation potential in the fetal liver (33), was reduced three-fold in CMPs but was increased significantly in GMPs. Although the CMP compartment was substantially reduced in Pbx1−/− embryos but increased in −77−/− fetal livers, both mutations similarly altered the CMP colony-forming potential, leading to reduced erythroid colonies and increased monocytes.

Bottom Line: The -77 established a vital sector of the myeloid progenitor transcriptome, conferring multipotentiality.Unlike the +9.5(-/-) embryos, hematopoietic stem cell genesis was unaffected in -77(-/-) embryos.These results illustrate a paradigm in which cis-elements in a locus differentially control stem and progenitor cell transitions, and therefore the individual cis-element alterations cause unique and overlapping disease phenotypes.

View Article: PubMed Central - PubMed

Affiliation: Carbone Cancer Center, Department of Cell and Regenerative Biology, University of Wisconsin School of Medicine and Public Health, Madison, WI 53705, USA. ; University of Wisconsin-Madison Blood Research Program, Madison, WI 53705, USA.

ABSTRACT
Cis-element encyclopedias provide information on phenotypic diversity and disease mechanisms. Although cis-element polymorphisms and mutations are instructive, deciphering function remains challenging. Mutation of an intronic GATA motif (+9.5) in GATA2, encoding a master regulator of hematopoiesis, underlies an immunodeficiency associated with myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML). Whereas an inversion relocalizes another GATA2 cis-element (-77) to the proto-oncogene EVI1, inducing EVI1 expression and AML, whether this reflects ectopic or physiological activity is unknown. We describe a mouse strain that decouples -77 function from proto-oncogene deregulation. The -77(-/-) mice exhibited a novel phenotypic constellation including late embryonic lethality and anemia. The -77 established a vital sector of the myeloid progenitor transcriptome, conferring multipotentiality. Unlike the +9.5(-/-) embryos, hematopoietic stem cell genesis was unaffected in -77(-/-) embryos. These results illustrate a paradigm in which cis-elements in a locus differentially control stem and progenitor cell transitions, and therefore the individual cis-element alterations cause unique and overlapping disease phenotypes.

No MeSH data available.


Related in: MedlinePlus