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Twist-2 controls myeloid lineage development and function.

Sharabi AB, Aldrich M, Sosic D, Olson EN, Friedman AD, Lee SH, Chen SY - PLoS Biol. (2008)

Bottom Line: Moreover, Twist-2 was found to have a contrasting effect on cytokine production: inhibiting the production of proinflammatory cytokines such as interleukin-12 (IL-12) and interferon-gamma (IFNgamma) while promoting the regulatory cytokine IL-10 by myeloid cells.In addition, Twist-2 was found to be essential for endotoxin tolerance.Thus, this study reveals the critical role of Twist-2 in regulating the development of myeloid lineages, as well as the function and inflammatory responses of mature myeloid cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology, Center for Cell and Gene Therapy, Baylor College of Medicine, Houston, Texas, USA.

ABSTRACT
Basic helix-loop-helix (bHLH) transcription factors play critical roles in lymphoid and erythroid development; however, little is known about their role in myeloid lineage development. In this study, we identify the bHLH transcription factor Twist-2 as a key negative regulator of myeloid lineage development, as manifested by marked increases in mature myeloid populations of macrophages, neutrophils, and basophils in Twist-2-deficient mice. Mechanistic studies demonstrate that Twist-2 inhibits the proliferation as well as differentiation of granulocyte macrophage progenitors (GMP) by interacting with and inhibiting the transcription factors Runx1 and C/EBPalpha. Moreover, Twist-2 was found to have a contrasting effect on cytokine production: inhibiting the production of proinflammatory cytokines such as interleukin-12 (IL-12) and interferon-gamma (IFNgamma) while promoting the regulatory cytokine IL-10 by myeloid cells. The data from further analyses suggest that Twist-2 activates the transcription factor c-Maf, leading to IL-10 expression. In addition, Twist-2 was found to be essential for endotoxin tolerance. Thus, this study reveals the critical role of Twist-2 in regulating the development of myeloid lineages, as well as the function and inflammatory responses of mature myeloid cells.

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Twist-2–Deficient Mice Have Significant Increases in Multiple Myeloid Cells in Multiple Organs(A and B) Semiquantitative (A) and quantitative (B) RT-PCR of sorted hematopoietic progenitor populations showing constitutive expression of Twist-2 in GMP and CMP populations. Data are normalized to 18s rRNA internal controls and representative of two independent experiments. Error bars indicate the standard deviation (SD).(C) Splenocytes of Twist-2 KO and WT littermates were analyzed by flow cytometry. Results are representative of eight independent pairs of mice analyzed. Splenic CD4+ and CD8+ T cells (upper panel), IgM+ and IgD+ B cells (middle panel), and CD11b+ and Gr-1+ granulocytes and macrophages (lower panel) are shown.(D) Flow cytometry showing expansion of CD11b+Gr-1+ myeloid cells in peripheral blood (upper panel), liver (middle panel), and BM (lower panel) of Twist-2 KO mice representative of six independent experiments.(E) Flow cytometric analysis of splenocytes showing the expansion of Gr-1lowCD11b+ F4/80high macrophages (upper panel) and F4/80lowCD11b+Gr-1high neutrophils (lower panel) in Twist-2 KO mice representative of three independent experiments.(F) Absolute cell counts of CD11c+ cells and two major DC subtypes, CD11c+CD11b+CD4+ and CD11c+CD11b−CD8+ cells, are both increased in Twist-2 KO spleen. Error bars indicate SD.(G) Images of whole bone mounts stained with H&E (left panels). BM touch preps stained with Wright-Giemsa (right panels) show higher numbers of mature myeloid cells in the Twist-2 KO BM.(H) Images of peripheral blood smears with Wright-Giemsa stain showing hypersegmented neutrophils and enlarged monocytoid cells in Twist-2 KO mice. Images are representative of four independent sets of peripheral blood smears. The arrow indicates the hypersegmented nuclei of the neutrophils.
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pbio-0060316-g001: Twist-2–Deficient Mice Have Significant Increases in Multiple Myeloid Cells in Multiple Organs(A and B) Semiquantitative (A) and quantitative (B) RT-PCR of sorted hematopoietic progenitor populations showing constitutive expression of Twist-2 in GMP and CMP populations. Data are normalized to 18s rRNA internal controls and representative of two independent experiments. Error bars indicate the standard deviation (SD).(C) Splenocytes of Twist-2 KO and WT littermates were analyzed by flow cytometry. Results are representative of eight independent pairs of mice analyzed. Splenic CD4+ and CD8+ T cells (upper panel), IgM+ and IgD+ B cells (middle panel), and CD11b+ and Gr-1+ granulocytes and macrophages (lower panel) are shown.(D) Flow cytometry showing expansion of CD11b+Gr-1+ myeloid cells in peripheral blood (upper panel), liver (middle panel), and BM (lower panel) of Twist-2 KO mice representative of six independent experiments.(E) Flow cytometric analysis of splenocytes showing the expansion of Gr-1lowCD11b+ F4/80high macrophages (upper panel) and F4/80lowCD11b+Gr-1high neutrophils (lower panel) in Twist-2 KO mice representative of three independent experiments.(F) Absolute cell counts of CD11c+ cells and two major DC subtypes, CD11c+CD11b+CD4+ and CD11c+CD11b−CD8+ cells, are both increased in Twist-2 KO spleen. Error bars indicate SD.(G) Images of whole bone mounts stained with H&E (left panels). BM touch preps stained with Wright-Giemsa (right panels) show higher numbers of mature myeloid cells in the Twist-2 KO BM.(H) Images of peripheral blood smears with Wright-Giemsa stain showing hypersegmented neutrophils and enlarged monocytoid cells in Twist-2 KO mice. Images are representative of four independent sets of peripheral blood smears. The arrow indicates the hypersegmented nuclei of the neutrophils.

Mentions: In order to identify bHLH factors that regulate myeloid development, we first used Gene Expression Omnibus (GEO) microarray dataset GSE3722 to analyze the expression of a panel of candidate genes, including known myeloid transcription factors and bHLH factors in CMP and GMP (Figure S1). Subsequent semiquantitative reverse transcription PCR (RT-PCR) analysis of sorted hematopoietic progenitors from wild-type (WT) C57B/6 mice (Figure S1) showed that Twist-2 was preferentially expressed in Lin−IL7R−Sca-1−c-Kit+CD34+FcγRII/IIIhigh GMP and Lin−IL7R−Sca-1−c-Kit+CD34+FcγRII/III− CMP, but not or weakly expressed in the Lin−IL7R−Sca-1+c-Kit+ hematopoietic progenitor compartment containing HSC or Lin−IL7R+Sca-1lowc-Kitlow common lymphoid progenitors (CLP) (Figure 1A). In contrast to the preferential expression of Twist-2, Twist-1 was expressed in all types of progenitors we examined (Figure 1A). Quantitative RT-PCR assays further confirmed the preferential and constitutive expression of Twist-2 in CMP and GMP populations (Figure 1B).


Twist-2 controls myeloid lineage development and function.

Sharabi AB, Aldrich M, Sosic D, Olson EN, Friedman AD, Lee SH, Chen SY - PLoS Biol. (2008)

Twist-2–Deficient Mice Have Significant Increases in Multiple Myeloid Cells in Multiple Organs(A and B) Semiquantitative (A) and quantitative (B) RT-PCR of sorted hematopoietic progenitor populations showing constitutive expression of Twist-2 in GMP and CMP populations. Data are normalized to 18s rRNA internal controls and representative of two independent experiments. Error bars indicate the standard deviation (SD).(C) Splenocytes of Twist-2 KO and WT littermates were analyzed by flow cytometry. Results are representative of eight independent pairs of mice analyzed. Splenic CD4+ and CD8+ T cells (upper panel), IgM+ and IgD+ B cells (middle panel), and CD11b+ and Gr-1+ granulocytes and macrophages (lower panel) are shown.(D) Flow cytometry showing expansion of CD11b+Gr-1+ myeloid cells in peripheral blood (upper panel), liver (middle panel), and BM (lower panel) of Twist-2 KO mice representative of six independent experiments.(E) Flow cytometric analysis of splenocytes showing the expansion of Gr-1lowCD11b+ F4/80high macrophages (upper panel) and F4/80lowCD11b+Gr-1high neutrophils (lower panel) in Twist-2 KO mice representative of three independent experiments.(F) Absolute cell counts of CD11c+ cells and two major DC subtypes, CD11c+CD11b+CD4+ and CD11c+CD11b−CD8+ cells, are both increased in Twist-2 KO spleen. Error bars indicate SD.(G) Images of whole bone mounts stained with H&E (left panels). BM touch preps stained with Wright-Giemsa (right panels) show higher numbers of mature myeloid cells in the Twist-2 KO BM.(H) Images of peripheral blood smears with Wright-Giemsa stain showing hypersegmented neutrophils and enlarged monocytoid cells in Twist-2 KO mice. Images are representative of four independent sets of peripheral blood smears. The arrow indicates the hypersegmented nuclei of the neutrophils.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC2602725&req=5

pbio-0060316-g001: Twist-2–Deficient Mice Have Significant Increases in Multiple Myeloid Cells in Multiple Organs(A and B) Semiquantitative (A) and quantitative (B) RT-PCR of sorted hematopoietic progenitor populations showing constitutive expression of Twist-2 in GMP and CMP populations. Data are normalized to 18s rRNA internal controls and representative of two independent experiments. Error bars indicate the standard deviation (SD).(C) Splenocytes of Twist-2 KO and WT littermates were analyzed by flow cytometry. Results are representative of eight independent pairs of mice analyzed. Splenic CD4+ and CD8+ T cells (upper panel), IgM+ and IgD+ B cells (middle panel), and CD11b+ and Gr-1+ granulocytes and macrophages (lower panel) are shown.(D) Flow cytometry showing expansion of CD11b+Gr-1+ myeloid cells in peripheral blood (upper panel), liver (middle panel), and BM (lower panel) of Twist-2 KO mice representative of six independent experiments.(E) Flow cytometric analysis of splenocytes showing the expansion of Gr-1lowCD11b+ F4/80high macrophages (upper panel) and F4/80lowCD11b+Gr-1high neutrophils (lower panel) in Twist-2 KO mice representative of three independent experiments.(F) Absolute cell counts of CD11c+ cells and two major DC subtypes, CD11c+CD11b+CD4+ and CD11c+CD11b−CD8+ cells, are both increased in Twist-2 KO spleen. Error bars indicate SD.(G) Images of whole bone mounts stained with H&E (left panels). BM touch preps stained with Wright-Giemsa (right panels) show higher numbers of mature myeloid cells in the Twist-2 KO BM.(H) Images of peripheral blood smears with Wright-Giemsa stain showing hypersegmented neutrophils and enlarged monocytoid cells in Twist-2 KO mice. Images are representative of four independent sets of peripheral blood smears. The arrow indicates the hypersegmented nuclei of the neutrophils.
Mentions: In order to identify bHLH factors that regulate myeloid development, we first used Gene Expression Omnibus (GEO) microarray dataset GSE3722 to analyze the expression of a panel of candidate genes, including known myeloid transcription factors and bHLH factors in CMP and GMP (Figure S1). Subsequent semiquantitative reverse transcription PCR (RT-PCR) analysis of sorted hematopoietic progenitors from wild-type (WT) C57B/6 mice (Figure S1) showed that Twist-2 was preferentially expressed in Lin−IL7R−Sca-1−c-Kit+CD34+FcγRII/IIIhigh GMP and Lin−IL7R−Sca-1−c-Kit+CD34+FcγRII/III− CMP, but not or weakly expressed in the Lin−IL7R−Sca-1+c-Kit+ hematopoietic progenitor compartment containing HSC or Lin−IL7R+Sca-1lowc-Kitlow common lymphoid progenitors (CLP) (Figure 1A). In contrast to the preferential expression of Twist-2, Twist-1 was expressed in all types of progenitors we examined (Figure 1A). Quantitative RT-PCR assays further confirmed the preferential and constitutive expression of Twist-2 in CMP and GMP populations (Figure 1B).

Bottom Line: Moreover, Twist-2 was found to have a contrasting effect on cytokine production: inhibiting the production of proinflammatory cytokines such as interleukin-12 (IL-12) and interferon-gamma (IFNgamma) while promoting the regulatory cytokine IL-10 by myeloid cells.In addition, Twist-2 was found to be essential for endotoxin tolerance.Thus, this study reveals the critical role of Twist-2 in regulating the development of myeloid lineages, as well as the function and inflammatory responses of mature myeloid cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology, Center for Cell and Gene Therapy, Baylor College of Medicine, Houston, Texas, USA.

ABSTRACT
Basic helix-loop-helix (bHLH) transcription factors play critical roles in lymphoid and erythroid development; however, little is known about their role in myeloid lineage development. In this study, we identify the bHLH transcription factor Twist-2 as a key negative regulator of myeloid lineage development, as manifested by marked increases in mature myeloid populations of macrophages, neutrophils, and basophils in Twist-2-deficient mice. Mechanistic studies demonstrate that Twist-2 inhibits the proliferation as well as differentiation of granulocyte macrophage progenitors (GMP) by interacting with and inhibiting the transcription factors Runx1 and C/EBPalpha. Moreover, Twist-2 was found to have a contrasting effect on cytokine production: inhibiting the production of proinflammatory cytokines such as interleukin-12 (IL-12) and interferon-gamma (IFNgamma) while promoting the regulatory cytokine IL-10 by myeloid cells. The data from further analyses suggest that Twist-2 activates the transcription factor c-Maf, leading to IL-10 expression. In addition, Twist-2 was found to be essential for endotoxin tolerance. Thus, this study reveals the critical role of Twist-2 in regulating the development of myeloid lineages, as well as the function and inflammatory responses of mature myeloid cells.

Show MeSH
Related in: MedlinePlus