Alternative Runx1 promoter usage in mouse developmental hematopoiesis

Bee, Thomas; Liddiard, Kate; Swiers, Gemma; Bickley, Sorrel R. B.; Vink, Chris S.; Jarratt, Andrew; Hughes, Jim R.; Medvinsky, Alexander; Bruijn, Marella F. T. R. de

doi:10.1016/j.bcmd.2009.03.011

Cited by 51 publications

(73 citation statements)

References 60 publications

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“…18,20,26,27 However, the physiologic role of P1-Runx1 and/or P2-Runx1 in the first definitive hematopoietic cells of the mouse embryo is not clear. On the one hand, the predominance of P1-Runx1 in FL hematopoietic stem and progenitor cells has been interpreted as the P1 being the critical promoter during HSC emergence.…”

Section: Introductionmentioning

confidence: 99%

“…20 Embryos were collected in phosphate-buffered saline (PBS; Gibco, Invitrogen) supplemented with 10% fetal calf serum (FCS; Biosera), 50 U/mL penicillin, and 50 g/mL streptomycin (Cambrex Corporation).…”

Section: Timed Matings and Embryo Collectionmentioning

confidence: 99%

“…14,15 Vertebrate Runx1 is transcribed from 2 alternative promoters, the distal P1 and proximal P2. 4,[16][17][18][19][20] Alternative Runx1 promoter usage results in the generation of a series of transcripts that differ in their untranslated regions and/or protein-coding exons, influencing mRNA stability, efficiency of translation, and Runx1 protein structure (Levanon and Groner, 4 Bee et al, 20 Pozner et al, 21 Ben-Ami et al, 22 and references therein). Interestingly, Runx1 isoforms were reported to add to the functional complexity of Runx1 in vitro and in discrete cellular processes in vivo.…”

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confidence: 99%

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Nonredundant roles for Runx1 alternative promoters reflect their activity at discrete stages of developmental hematopoiesis

Bee

Swiers

Muroi³

et al. 2010

Blood

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The transcription factor Runx1 is a pivotal regulator of definitive hematopoiesis in mouse ontogeny. Vertebrate Runx1 is transcribed from 2 promoters, the distal P1 and proximal P2, which provide a paradigm of the complex transcriptional and translational control of Runx1 function. However, very little is known about the biologic relevance of alternative Runx1 promoter usage in definitive hematopoietic cell emergence. Here we report that both promoters are active at the very onset of definitive hematopoiesis, with a skewing toward the P2. Moreover, functional and morphologic analysis of a novel P1-null and an attenuated P2 mouse model revealed that although both promoters play important nonredundant roles in the emergence of definitive hematopoietic cells, the proximal P2 was most critically required for this. The nature of the observed phenotypes is indicative of a differential contribution of the P1 and P2 promoters to the control of overall Runx1 levels, where and when this is most critically required. In addition, the dynamic expression of P1-Runx1 and P2-Runx1 points at a requirement for Runx1 early in development, when the P2 is still the prevalent promoter in the emerging hemogenic endothelium and/or first committed hematopoietic cells. (Blood. 2010;115(15): 3042-3050) IntroductionThe generation of the definitive hematopoietic system during embryogenesis critically depends on the transcription factor Runx1. In mice, homozygous loss of Runx1 function results in embryonic lethality attributable to a complete lack of functional definitive hematopoietic stem cells (HSCs) and progenitor cells and hemorrhages in the central nervous system. 1-3 Runx1 belongs to the family of runt-domain transcription factors. The 3 mammalian members of this family, Runx1, 2, and 3, all are important developmental regulators and bind to the same DNA motif. 4 Although both Runx2 and Runx3 have been implicated in hematopoiesis, only Runx1 has a role in the emergence of definitive hematopoietic cells, 5 reflecting its specific expression at hemogenic sites. 6,7 Recently, it was shown that Runx1 is required in VE-cadherin ϩ cells of the embryo, within the developmental window that starts with the initiation of Runx1 expression in these cells and ends when/before definitive HSCs reach the embryonic day (E) 11 fetal liver (FL). 8 Although the precise developmental stage(s) at which Runx1 is required within this window remains to be determined, it is generally believed to be at the transition of hemogenic endothelium to definitive hematopoietic cells. 6,[8][9][10] In the adult, Runx1 is no longer critically required in HSCs, although it still plays important roles in maintaining hematopoietic homeostasis and in the generation of specific hematopoietic cells/lineages. [11][12][13] Not only the expression pattern of Runx1 but also its levels need to be tightly controlled for the normal emergence of HSCs in the embryo. 3 To gain insight into how this is achieved, we have initiated studies into the transcriptional regulation of Runx1. 14,15...

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Section: Introductionmentioning

confidence: 99%

Section: Timed Matings and Embryo Collectionmentioning

confidence: 99%

mentioning

confidence: 99%

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Nonredundant roles for Runx1 alternative promoters reflect their activity at discrete stages of developmental hematopoiesis

Bee

Swiers

Muroi³

et al. 2010

Blood

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“…Mutation of RUNX1 Serines 48, 303, or 424 to Aspartic Acid Reduces HDAC1 or HDAC3 Interaction-We next sought to test the idea that cdk phosphorylation of RUNX1 leads to reduced HDAC interaction, focusing on HDAC1 and HDAC3, for which RUNX1 has the highest affinity (21), and on the 480 residue RUNX1c isoform, which dominates during adult hematopoiesis (23). Myc-tagged RUNX1, RUNX1(tripleA), or RUNX1(tripleD) were co-expressed in 293T cells along with FLAG-tagged HDAC1 or HDAC3, followed by immunoprecipitation with Myc antibody or Ig control and then Western blotting with FLAG or Myc antibodies.…”

Section: Resultsmentioning

confidence: 99%

Phosphorylation of RUNX1 by Cyclin-dependent Kinase Reduces Direct Interaction with HDAC1 and HDAC3

Guo

Friedman

2011

Journal of Biological Chemistry

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RUNX1 regulates formation of the definitive hematopoietic stem cell and its subsequent lineage maturation, and mutations of RUNX1 contribute to leukemic transformation. Phosphorylation of Ser-48, Ser-303, and Ser-424 by cyclin-dependent kinases (cdks) increases RUNX1 trans-activation activity without perturbing p300 interaction. We now find that endogenous RUNX1 interacts with endogenous HDAC1 or HDAC3. Mutation of the three RUNX1 serines to aspartic acid reduces co-immunoprecipitation with HDAC1 or HDAC3 when expressed in 293T cells; mutation of these three serines to alanine increases HDAC interaction, and mutation of each serine individually to aspartic acid also reduces these interactions. GST-RUNX1 isolated from bacterial extracts bound in vitro translated HDAC1 or HDAC3, and these interactions were weakened by mutation of Ser-48, Ser-303, and Ser-424 to aspartic acid. The ability of RUNX1 phosphorylation and not only serine to aspartic acid conversion to reduce HDAC1 binding was demonstrated using wild-type GST-RUNX1 phosphorylated in vitro using cdk1/cyclinB and by exposure of 293T cells transduced with RUNX1 and HDAC1 to roscovitine, a cdk inhibitor. Finally, RUNX1 or RUNX1(tripleD), in which Ser-48, Ser-303, and Ser-424 are mutated to aspartic acid, stimulated proliferation of transduced, lineage-negative murine marrow progenitors more potently than did RUNX1(tripleA), in which these serines are mutated to alanine, suggesting that stimulation of RUNX1 trans-activation by cdk-mediated reduction in HDAC interaction increases marrow progenitor cell proliferation. RUNX1 and its heterodimeric partner CBF␤ direct emergence of adult hematopoietic stem cells (HSC)2 from hemogenic endothelium during embryogenesis and participate in subsequent lymphoid, myeloid, or megakaryocyte lineage maturation in adult marrow (1-3). RUNX1 is commonly mutated or inhibited in acute myeloid or lymphoid leukemias, and RUNX1 is overexpressed as well in a subset of acute lymphoblastic leukemias (4, 5).RUNX1 also directly regulates G 1 to S cell cycle progression. The myeloid oncoproteins CBF␤-SMMHC or RUNX1-ETO dominantly inhibit RUNX1 activities and slow G 1 progression in hematopoietic cell lines or in murine or human marrow progenitors (6 -9). cdk4, cyclin D2, or c-Myc overcome inhibition of proliferation by these CBF oncoproteins (8, 10, 11); exogenous RUNX1 stimulates G 1 progression in 32Dcl3 or Ba/F3 cells (9, 10, 12), and stimulation of G 1 via deletion of the p16INK4a/p19ARF locus or expression of the viral E7 protein (which inactivates retinoblastoma protein) cooperates with CBF␤-SMMHC or TEL-RUNX1 to induce acute leukemia (13,14). Induction of cdk4 or cyclin D3 transcription may underlie stimulation of G 1 progression by RUNX1 (10,15).Regulation of cell proliferation by RUNX proteins represents an evolutionarily conserved activity. In the sea urchin Strongylocentrotus purpuratus, depletion of the RUNX ortholog SpRunt-1 reduces blastocyst cell proliferation and inhibits expression of cyclinD, whose promoter binds SpRunt-1...

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“…This 5-kb fragment is located 3.8 kb away from the reported RUNX1 promoter. 33 After cotransfection into leukemia cells, this 5-kb fragment drove the expression of the luciferase reporter gene to a greater degree than did the promoter-less basic vector (Fig. 1e).…”

Section: Identification Of a Novel Intragenic Long Noncoding Rna In Tmentioning

confidence: 93%

An intragenic long noncoding RNA interacts epigenetically with theRUNX1promoter and enhancer chromatin DNA in hematopoietic malignancies

Wang

Guo

et al. 2014

Int. J. Cancer

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RUNX1, a master regulator of hematopoiesis, is the most commonly perturbed target of chromosomal abnormalities in hematopoietic malignancies. The t(8;21) translocation is found in 30-40% of cases of acute myeloid leukemia (AML). Recent wholeexome sequencing also reveals mutations and deletions of RUNX1 in some solid tumors. We describe a RUNX1-intragenic long noncoding RNA RUNXOR that is transcribed as unspliced transcript from an upstream overlapping promoter. RUNXOR was upregulated in AML samples and in response to Ara-C treatment in vitro. RUNXOR utilizes its 3 0 -terminal fragment to directly interact with the RUNX1 promoter and enhancers and participates in the orchestration of an intrachromosomal loop. The 3 0 region of RUNXOR also participates in long-range interchromosomal interactions with chromatin regions that are involved in multiple RUNX1 translocations. These data suggest that RUNXOR noncoding RNA may function as a previously unidentified candidate component that is involved in chromosomal translocation in hematopoietic malignancies.

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Alternative Runx1 promoter usage in mouse developmental hematopoiesis

Cited by 51 publications

References 60 publications

Nonredundant roles for Runx1 alternative promoters reflect their activity at discrete stages of developmental hematopoiesis

Nonredundant roles for Runx1 alternative promoters reflect their activity at discrete stages of developmental hematopoiesis

Phosphorylation of RUNX1 by Cyclin-dependent Kinase Reduces Direct Interaction with HDAC1 and HDAC3

An intragenic long noncoding RNA interacts epigenetically with theRUNX1promoter and enhancer chromatin DNA in hematopoietic malignancies

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