scholarly journals PU.1 Enforces Hematopoietic Stem Cell Quiescence during Chronic Inflammation

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 822-822
Author(s):  
James Chavez ◽  
Jennifer L Rabe ◽  
Kelly Higa ◽  
Dirk Loeffler ◽  
Ahmed Nouraiz ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Protein Synthesis ◽  
Stem Cell ◽  
Chronic Inflammation ◽  
Hematopoietic Stem Cell ◽  
Synthesis Rate ◽  
Rna Seq ◽  
Hematopoietic Stem ◽  
Cell Cycle Entry ◽  
Insight Into

Hematopoietic stem cell (HSC) quiescence is crucial for maintaining lifelong blood production and preventing potentially toxic overproduction of blood cells. HSC are capable of re-entering quiescence following exposure to pro-inflammatory stimuli such as interleukin (IL)-1, implying the existence of one or more 'braking' mechanisms that limit and/or overcome the mitogenic properties of these signals to limit HSC cell cycle entry. However, mechanism(s) regulating HSC quiescence during chronic inflammation have yet to be fully elucidated. In the present study, we find that the master myeloid transcription factor PU.1 represses Myc-regulated cell cycle and protein synthesis pathways in HSC during chronic inflammation, thereby guarding HSC quiescence in this context. To gain insight into HSC cell cycle regulation in the context of chronic inflammatory signaling, we performed cell cycle and RNA-seq analysis on purified HSC from mice injected daily for 20 days with IL-1β. Strikingly, HSC from IL-1-treated mice remained in a quiescent state, and our RNA-seq analyses uncovered a significant decrease in mRNA transcripts from pathways related to cellular proliferation, translation, and ribosome biogenesis. Also, flow cytometry analyses revealed that IL-1 significantly reduced Myc protein expression and ribosomal protein S6 phosphorylation in HSC. Altogether, these data identify an inflammation-induced cell cycle restriction gene program that limits HSC proliferation in response to inflammation. Strikingly, ingenuity pathway analysis (IPA) predicted PU.1as a possible driver of the cell cycle restriction gene program. Indeed, PU.1 mRNA and protein levels in HSC increased significantly in PU.1-eYFP knockin reporter mice treated with IL-1. Furthermore, fractionating HSC based on reporter levels showed that IL-1-exposed HSC expressing high PU.1 levels activated the cell cycle restriction program more robustly than HSC with lower PU.1 levels or from untreated control mice. Along these lines, cell cycle analyses showed that PU.1-high HSC were quiescent following IL-1 exposure. Surprisingly, functional assays revealed that the PU.1-high fraction became enriched for functional HSC following IL-1 exposure, in contrast to a primarily PU.1-low phenotype under control conditions. Collectively, these data show that low Myc levels, a quiescent cell cycle state and functional HSC identity are all associated with high PU.1 levels during chronic inflammation. To address the requirement for PU.1, we compared HSC following IL-1 exposure from wild-type (WT) and PU.1-knockin (KI) mice, which express ~30% of normal PU.1 levels due to a point mutation in the PU.1 upstream regulatory element (URE). Strikingly, IL-1 induced aberrant HSC expansion in the BM of PU.1 KI mice, underwritten by IL-1-induced quiescence loss in PU.1 KI HSC. Excess cell cycle activity in HSC from IL-1-treated PU.1 KI mice was associated with Myc overexpression, hyperinduction of cycle and protein synthesis genes, and an increased protein synthesis rate. Interestingly, IL-1-treated PU.1 KI mice also exhibited exuberant platelet production and aberrant activation of megakaryocyte (Mk) lineage programs in HSC. Taken together, these data show PU.1 is required to limit HSC proliferation and cell cycle activity, and that failure to upregulate PU.1 during chronic inflammation leads to aberrant HSC expansion and dysregulated Mk lineage output. Altogether, our data identify PU.1 as a key enforcer of HSC dormancy, and as a regulator of HSC proliferation and proteostasis during chronic inflammation. As PU.1 has been shown to slow cell cycle entry to promote myeloid differentiation in actively proliferating cells, our results suggest a mechanistic conservation in which inflammation induces PU.1 expression and engages a similar cell cycle restriction program that preserves HSC quiescence. Such a mechanism can prevent excessive Mk lineage output leading to thrombosis, and/or HSC exhaustion. Importantly, these findings re-cast inflammation-induced PU.1 expression as a guardian of blood system function, rather than a pathogenic by-product. As PU.1 serves a tumor suppressor role, these findings also may provide insight into the link between inflammation and leukemogenesis, particularly in contexts where PU.1 function is impaired in leukemic stem cells (LSC) due to mutation or indirect mechanism(s). Disclosures No relevant conflicts of interest to declare.

scholarly journals GATA-3 regulates hematopoietic stem cell maintenance and cell-cycle entry

Blood ◽  
2012 ◽  
Vol 119 (10) ◽  
pp. 2242-2251 ◽  
Author(s):  
Chia-Jui Ku ◽  
Tomonori Hosoya ◽  
Ivan Maillard ◽  
James Douglas Engel
Keyword(s):  
Cell Cycle ◽  
Stem Cell ◽  
Hematopoietic Stem Cell ◽  
Hematopoietic Stem ◽  
Stem Cell Maintenance ◽  
Normal Number ◽  
Cell Cycle Entry ◽  
Null Mutant Mice ◽  
Th2 Differentiation

Abstract Maintaining hematopoietic stem cell (HSC) quiescence is a critical property for the life-long generation of blood cells. Approximately 75% of cells in a highly enriched long-term repopulating HSC (LT-HSC) pool (Lin−Sca1+c-KithiCD150+CD48−) are quiescent, with only a small percentage of the LT-HSCs in cycle. Transcription factor GATA-3 is known to be vital for the development of T cells at multiple stages in the thymus and for Th2 differentiation in the peripheral organs. Although it is well documented that GATA-3 is expressed in HSCs, a role for GATA-3 in any prethymic progenitor cell has not been established. In the present study, we show that Gata3-null mutant mice generate fewer LT-HSCs and that fewer Gata3-null LT-HSCs are in cycle. Furthermore, Gata3 mutant hematopoietic progenitor cells fail to be recruited into an increased cycling state after 5-fluorouracil–induced myelosuppression. Therefore, GATA-3 is required for the maintenance of a normal number of LT-HSCs and for their entry into the cell cycle.

scholarly journals Mitochondrial Morphology Controls Hematopoietic Stem Cell (HSC) Self-Renewal and Confers HSC Divisional Memory

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. SCI-20-SCI-20
Author(s):  
Marie-Dominique Filippi
Keyword(s):  
Cell Cycle ◽  
Stem Cell ◽  
Hematopoietic Stem Cell ◽  
Mitochondrial Morphology ◽  
Regenerative Capacity ◽  
Self Renewal ◽  
Principal Mechanism ◽  
Hematopoietic Stem ◽  
Replicative Stress ◽  
Cell Cycle Entry

Abstract Adult hematopoietic stem cell (HSC) have extensive regenerative capacity; but recent evidence strongly indicate that their actual capacity for self-renewal is limited. HSC attrition occurs with cumulative divisions and after acute stress, causing reduction in regenerative capacity of the progeny. The principal mechanism driving the loss of HSC potency with stress hematopoiesis is not completely understood. We will show and discuss the concept that regulatory programs that are activated with HSC activation and cell cycle entry cause a permanent remodeling of mitochondria. Mitochondria do not return to homeostasis after replicative stress despite HSC re-entry into quiescence and termination of the metabolic programs that were activated with HSC cell cycle entry. HSC keep dysfunctional mitochondria after replicative stress and this loss in mitochondrial fidelity drives the decline of the HSC pool after regenerative stress. The mitochondrial remodeling with replicative stress is at least in part due to a loss in mitochondrial quality control, notably Drp1-mediated mitochondrial dynamics. Further, single cell RNA-seq data indicate that HSC carrying dysfunctional mitochondria fail to synchronize the transcriptional control of core cell cycle and metabolic components in subsequent division providing unexpected mechanism of why HSC that have divided are less potent than HSC that have never divided in similar microenvironment. Thus, the loss of fidelity of mitochondria homeostasis drives HSC attrition and serves as one source of cellular memory of hematopoietic stem cell replicative history. Disclosures No relevant conflicts of interest to declare.

3014 – ORAL PRESENTATION: SINGLE-CELL RNA-SEQ REVEALS A CONCOMITANT DELAY IN DIFFERENTIATION AND CELL CYCLE OF AGED HEMATOPOIETIC STEM CELL

2020 ◽  
Vol 88 ◽  
pp. S42
Author(s):  
Léonard Hérault ◽  
Mathilde Poplineau ◽  
Adrien Mazuel ◽  
Nadine Platet ◽  
Élisabeth Remy ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Stem Cell ◽  
Single Cell ◽  
Hematopoietic Stem Cell ◽  
Oral Presentation ◽  
Rna Seq ◽  
Hematopoietic Stem

scholarly journals PU.1 enforces quiescence and limits hematopoietic stem cell expansion during inflammatory stress

2020 ◽  
Author(s):  
James S. Chavez ◽  
Jennifer L. Rabe ◽  
Dirk Loeffler ◽  
Kelly C. Higa ◽  
Giovanny Hernandez ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Protein Synthesis ◽  
Stem Cell ◽  
Hematopoietic Stem Cell ◽  
Hematological Malignancy ◽  
Hematopoietic Stem ◽  
Inflammatory Stress ◽  
Stem Cell Expansion ◽  
Cell Cycle Genes ◽  
Hematopoietic Stem Cell Expansion

SummaryLoss of hematopoietic stem cell (HSC) quiescence and resulting clonal expansion are common initiating events in the development of hematological malignancy. Likewise, chronic inflammation related to aging, disease and/or tissue damage is associated with leukemia progression, though its role in oncogenesis is not clearly defined. Here, we show that PU.1-dependent repression of protein synthesis and cell cycle genes in HSC enforces homeostatic protein synthesis levels and HSC quiescence in response to IL-1 stimulation. These genes are constitutively de-repressed in PU.1-deficient HSC, leading to activation of protein synthesis, loss of quiescence and aberrant expansion of HSC. Taken together, our data identify a mechanism whereby HSC regulate their cell cycle activity and pool size in response to chronic inflammatory stress.

miR-146a regulates hematopoietic stem cell maintenance and cell cycle entry

2013 ◽  
Vol 41 (8) ◽  
pp. S17
Author(s):  
Joanna Wegrzyn Woltosz ◽  
David Knapp ◽  
Michael Copley ◽  
Rawa Ibrahim ◽  
Patricia Umlandt ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Stem Cell ◽  
Hematopoietic Stem Cell ◽  
Hematopoietic Stem ◽  
Stem Cell Maintenance ◽  
Cell Cycle Entry ◽  
Cell Maintenance

scholarly journals Hematopoietic Stem Cell Function in a Murine Model of Sickle Cell Disease

Anemia ◽  
2012 ◽  
Vol 2012 ◽  
pp. 1-9 ◽  
Author(s):  
Elisabeth H. Javazon ◽  
Mohamed Radhi ◽  
Bagirath Gangadharan ◽  
Jennifer Perry ◽  
David R. Archer
Keyword(s):  
Cell Cycle ◽  
Lipid Peroxidation ◽  
Bone Marrow ◽  
Stem Cell ◽  
Sickle Cell Disease ◽  
Hematopoietic Stem Cell ◽  
Cell Function ◽  
Cell Disease ◽  
Hematopoietic Progenitor ◽  
Hematopoietic Stem

Previous studies have shown that the sickle environment is highly enriched for reactive oxygen species (ROS). We examined the oxidative effects of sickle cell disease on hematopoietic stem cell function in a sickle mouse model.In vitrocolony-forming assays showed a significant decrease in progenitor colony formation derived from sickle compared to control bone marrow (BM). Sickle BM possessed a significant decrease in the KSL (c-kit+, Sca-1+, Lineage−) progenitor population, and cell cycle analysis showed that there were fewer KSL cells in the G0phase of the cell cycle compared to controls. We found a significant increase in both lipid peroxidation and ROS in sickle-derived KSL cells.In vivoanalysis demonstrated that normal bone marrow cells engraft with increased frequency into sickle mice compared to control mice. Hematopoietic progenitor cells derived from sickle mice, however, demonstrated significant impairment in engraftment potential. We observed partial restoration of engraftment by n-acetyl cysteine (NAC) treatment of KSL cells prior to transplantation. Increased intracellular ROS and lipid peroxidation combined with improvement in engraftment following NAC treatment suggests that an altered redox environment in sickle mice affects hematopoietic progenitor and stem cell function.

ASXL2 Is a Novel Mediator of RUNX1-ETO Transcriptional Function and Collaborates with RUNX1-ETO to Promote Leukemogenesis

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 302-302
Author(s):  
Jean-Baptiste Micol ◽  
Nicolas Duployez ◽  
Alessandro Pastore ◽  
Robert Williams ◽  
Eunhee Kim ◽  
...  
Keyword(s):  
Stem Cell ◽  
Cell Line ◽  
Hematopoietic Stem Cell ◽  
Binding Sites ◽  
Target Genes ◽  
Cell Function ◽  
Rna Seq ◽  
Hematopoietic Stem

Abstract Mutations in Addition of Sex Combs Like 1 (ASXL1) are common in patients with myeloid leukemias. More recently, mutations in ASXL2, a paralog of ASXL1 with ~40% shared amino acid homology, have been discovered to occur specifically in patients with acute myeloid leukemia (AML) patients bearing the RUNX1-ETO (AML1-ETO; RUNX1-RUNX1T1) translocation and are amongst the most common mutations in RUNX1-ETO AML (mutated in 20-25% of patients). Although ASXL1 is critical for Polycomb Repressive Complex 2 function in myeloid hematopoietic cells and loss of Asxl1 recapitulates key aspects of myelodysplastic syndrome (MDS), the function of ASXL2 in normal or malignant hematopoiesis is unknown. We therefore set out to perform a functional comparison of ASXL1and ASXL2on hematopoiesis and transcription and determine the functional basis for frequent mutations in RUNX1-ETO AML. In vitro analyses of ASXL2 insertion/deletion mutations revealed that these mutations resulted in substantial reduction of ASXL2 protein expression, stability, and half-life. We therefore generated Asxl2 conditional knockout (cKO) mice to delineate the effect of ASXL2 loss on hematopoiesis. Competitive (Fig. 1A) and noncompetitive transplantation revealed that Asxl2 or compound Asxl1/2 loss resulted in cell-autonomous, rapid defects of hematopoietic stem cell function, self-renewal, and number with peripheral blood leukopenia and thrombocytopenia but without any obvious MDS features- phenotypes distinct from Asxl1 cKO mice. Mice with heterozygous deletion of Asxl2 demonstrated an intermediate phenotype between control and homozygous cKO mice indicating a gene dosage effect of Asxl2 loss. RNA sequencing (RNA-seq) of hematopoietic stem/progenitor cells from Asxl2- and Asxl1-deficient mice revealed twenty-fold greater differentially expressed genes in Asxl2 cKO mice relative to Asxl1 cKO mice. Interestingly, genes differentially expressed with Asxl2 loss significantly overlapped with direct transcriptional targets of RUNX1-ETO, findings not seen in Asxl1 cKO mice (Fig. 1B). Asxl2 target genes appeared to also be targets of RUNX1, a key gene repressed by RUNX1-ETO to promote leukemogenesis. Consistent with this, genome-wide analysis of Asxl2 binding sites through anti-Asxl2 ChIP-seq revealed that Asxl2 binding sites substantially overlap with those of Runx1. Overall, the above data suggest that Asxl2 may be a critical mediator of RUNX1-ETO mediated leukemogenesis by affecting the expression of RUNX1 and/or RUNX1-ETO target genes. RNA-seq of primary RUNX1-ETO AML patient samples revealed that ASXL2-mutant RUNX1-ETO patients form a distinct transcriptional subset of RUNX1-ETO AML (Fig. 1C) suggesting a specific role of ASXL2 in leukemogenesis. To functionally interrogate the role of ASXL2 loss in RUNX1-ETO mediated leukemogenesis we first utilized an in vitro model with RNAi-mediated depletion of ASXL1 or ASXL2 in the SKNO1 cell line (the only ASXL-wildtype human RUNX1-ETO cell line). RNA-seq revealed distinct target genes dysregulated by ASXL1 versus ASXL2 loss in these cells without any significant overlap. Anti-ASXL2, RUNX1, and RUNX1-ETO ChIPSeq in SKNO1 cells revealed significant co-occupancy of ASXL2 with RUNX1 and RUNX1-ETO binding sites. Moreover, analysis of histone modification ChIPSeq revealed an enrichment in intergenic and enhancer H3K4me1 abundance following ASXL2 loss in SKNO1 cells. Next, to understand the in vivo effects of Asxl2 loss in the context of RUNX1-ETO, we performed retroviral bone marrow (BM) transplantation assays using RUNX1-ETO9a in Asxl2 cKO mice. In contrast to the failure of hematopoietic stem cell function with Asxl2 deletion alone, mice reconstituted with BM cells expressing RUNX1-ETO9a in Asxl2-deficient background had a shortened leukemia-free survival compared to Asxl2 -wildtype control. Overall, these data reveal that ASXL2 is required for hematopoiesis and has differing biological and transcriptional functions from ASXL1. Moreover, this work identifies ASXL2 as a novel mediator of RUNX1-ETOtranscriptional function and provides a new model of penetrant RUNX1-ETO AML based on genetic events found in a substantial proportion of t(8;21) AML patients. Further interrogation of the enhancer alterations generated by ASXL2 loss in RUNX1-ETO AML may highlight new therapeutic approaches for this subset of AML. Figure 1. Figure 1. Disclosures No relevant conflicts of interest to declare.

Sign in / Sign up

Export Citation Format

Share Document