scholarly journals Musashi-2 controls cell fate, lineage bias, and TGF-β signaling in HSCs

2014 ◽  
Vol 211 (1) ◽  
pp. 71-87 ◽  
Author(s):  
Sun-Mi Park ◽  
Raquel P. Deering ◽  
Yuheng Lu ◽  
Patrick Tivnan ◽  
Steve Lianoglou ◽  
...  
Keyword(s):  
Cell Fate ◽  
Rna Binding ◽  
Asymmetric Cell Division ◽  
Cell Function ◽  
Rna Binding Protein ◽  
Transcriptome Profiling ◽  
Hematopoietic Stem ◽  
Rna Translation ◽  
Important Regulator ◽  
Global Transcriptome

Hematopoietic stem cells (HSCs) are maintained through the regulation of symmetric and asymmetric cell division. We report that conditional ablation of the RNA-binding protein Msi2 results in a failure of HSC maintenance and engraftment caused by a loss of quiescence and increased commitment divisions. Contrary to previous studies, we found that these phenotypes were independent of Numb. Global transcriptome profiling and RNA target analysis uncovered Msi2 interactions at multiple nodes within pathways that govern RNA translation, stem cell function, and TGF-β signaling. Msi2-null HSCs are insensitive to TGF-β–mediated expansion and have decreased signaling output, resulting in a loss of myeloid-restricted HSCs and myeloid reconstitution. Thus, Msi2 is an important regulator of the HSC translatome and balances HSC homeostasis and lineage bias.

Direct RNA Target Analysis Identifies Signatures That Define Msi2's Role in Sustaining HSC Cell Fate.

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 2324-2324
Author(s):  
Sun Mi Park ◽  
Raquel Deering ◽  
Yuheng Lu ◽  
Patrick Tivnan ◽  
Varun Shenoy ◽  
...  
Keyword(s):  
Stem Cell ◽  
Cell Division ◽  
Notch Signaling ◽  
Cell Fate ◽  
Rna Binding ◽  
Rna Binding Protein ◽  
Self Renewal ◽  
Hematopoietic Stem ◽  
Gene Set

Abstract Abstract 2324 Hematopoietic stem cells (HSC) must maintain normal cell fate decisions between symmetric and asymmetric divisions as alterations can lead to hematopoietic malignancies. The MSI2 RNA binding protein is upregulated in patients with a poor clinical prognosis in acute myeloid leukemia and in the blast crisis phase of chronic myelogenous leukemia. The related RNA-binding protein Msi1 has been shown to block translation of specific target mRNAs by interacting with the 3'UTR. To understand the role of Msi2 in both normal and leukemic contexts, we are characterizing mice with a conditional deletion of Msi2 in the hematopoietic compartment. Msi2 conditional knockouts have reduced overall number of HSCs in the bone marrow and have decreased engraftment capacity in congenic transplants, however the molecular mechanisms through which Msi2 elicits these effects remains unclear. To understand how Msi2 alters HSC self renewal, we utilized Mx1-Cre::Msi2flox/flox conditional mice, enabling Msi2 inactivation via poly(I):poly(C) injection. We first tested if Msi2 deleted HSCs or MPPs had an intrinsic defect in cell fate decision and proliferation. We performed proliferation and colony assays on sorted HSCs and MPPs and found reduced cellular numbers in both the HSCs and MPPs with an increase in the frequency of more differentiated cells based on Mac/Gr1 staining after seven days. These reduced overall cell numbers in vitro may have resulted from a defect in maintaining the stem cell population or a decrease in proliferative capacity. To assess if there was a defect in the initial HSC cell division, we examined Numb protein levels and distribution as a surrogate readout for asymmetric cell division. Numb mRNA is a known target of Msi binding and translational repression. Surprisingly, in the Msi2 null HSC or MPPs we observed no difference in the overall staining of Numb indicating that Msi2 deficiency does not globally alter Numb levels. However, we did observe a decrease in the percentage of cells that underwent asymmetric Numb segregation in the MPPs and an increase in the percentage of cells that showed increased Numb staining in some of the daughter pairs in both HSCs and MPPs, indicating increased commitment away from the hematopoietic stem and progenitors. Although controversial, Notch signaling has been implicated in self renewal of HSCs and as a critical downstream target of the Msi family through Numb inactivation. However, the Notch signaling pathway was not noted to be significantly altered in gene set analysis from microarrays performed on Msi2 deleted HSC enriched populations. Moreover using quantitative PCR for Notch target genes in sorted Linlow, c-kit+ and Sca+(LSK) cells we found no statistical difference in the expression of Notch targets (Notch1, Notch2, Hes1 or Myc). These interesting negative results prompted us take a more global approach in characterizing the direct targets of Msi2 in hematopoietic cells. Due to the requirement for a large number of cells, we utilized K562 cells overexpressing a FLAG-tagged version of MSI2 to identify its direct RNA-binding targets. Using recently developed technologies, we performed experiments with UV-cross-linked and immunoprecipitated MSI2 which was then followed by RNA-sequencing (HITS-CLIP) to identify the global direct binding. Our analysis characterized the distribution of binding across the genome. Additionally, gene set enrichment analysis (GSEA) indicated a positive correlation of genes that were upregulated in the Msi2 deleted LSKs and the top 3-fold bound RNA targets (2,713 genes). Using the entire RNA target list we identified gene set signatures including “Cell Cycle”, “Self-renewal” and “HSC to CMP” that matched our results in the microarray from Msi2 deleted LSKs. In summary, direct RNA target analysis for MSI2 in human leukemia cells overlapped with self renewal and differentiation gene sets in mouse HSC enriched populations and correlated with the phenotypes we observed in isolated HSCs grown in vitro lacking Msi2. These results uncover the complexity of MSI2 RNA binding targets and have important implications for both normal and leukemic stem cell biology. Disclosures: Ebert: Celgene: Consultancy; Genoptix: Consultancy.

Post-transcriptional regulation in hematopoiesis: RNA binding proteins take control

2019 ◽  
Vol 97 (1) ◽  
pp. 10-20 ◽  
Author(s):  
Laura P.M.H. de Rooij ◽  
Derek C.H. Chan ◽  
Ava Keyvani Chahi ◽  
Kristin J. Hope
Keyword(s):  
Transcriptional Regulation ◽  
Stem Cell ◽  
Cell Fate ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Control Of Gene Expression ◽  
Hematopoietic Stem ◽  
Cell Fate Decisions ◽  
Post Transcriptional Regulation

Normal hematopoiesis is sustained through a carefully orchestrated balance between hematopoietic stem cell (HSC) self-renewal and differentiation. The functional importance of this axis is underscored by the severity of disease phenotypes initiated by abnormal HSC function, including myelodysplastic syndromes and hematopoietic malignancies. Major advances in the understanding of transcriptional regulation of primitive hematopoietic cells have been achieved; however, the post-transcriptional regulatory layer that may impinge on their behavior remains underexplored by comparison. Key players at this level include RNA-binding proteins (RBPs), which execute precise and highly coordinated control of gene expression through modulation of RNA properties that include its splicing, polyadenylation, localization, degradation, or translation. With the recent identification of RBPs having essential roles in regulating proliferation and cell fate decisions in other systems, there has been an increasing appreciation of the importance of post-transcriptional control at the stem cell level. Here we discuss our current understanding of RBP-driven post-transcriptional regulation in HSCs, its implications for normal, perturbed, and malignant hematopoiesis, and the most recent technological innovations aimed at RBP–RNA network characterization at the systems level. Emerging evidence highlights RBP-driven control as an underappreciated feature of primitive hematopoiesis, the greater understanding of which has important clinical implications.

scholarly journals TWIST1 Preserves Hematopoietic Stem Cell Function Via Repression of Cacna1b Expression

Blood ◽  
2020 ◽  
Vol 136 (Supplement 1) ◽  
pp. 12-12
Author(s):  
Nan Wang ◽  
Jing Yin ◽  
Na You ◽  
Dan Guo ◽  
Yangyang Zhao ◽  
...  
Keyword(s):  
Stem Cell ◽  
Steady State ◽  
Calcium Channel ◽  
Cell Fate ◽  
Hematopoietic Stem Cell ◽  
Cell Function ◽  
Conflicts Of Interest ◽  
Hematopoietic Stem ◽  
Voltage Gated ◽  
Voltage Gated Calcium Channel

The mitochondria of hematopoietic stem cell (HSC) play crucial roles in regulating cell fate and in preserving HSC functionality and survival. However, the mechanism underlying its regulation remain poorly understood. Here, we identify transcription factor TWIST1 as a novel regulator of HSC maintenance through modulating mitochondrial function. We demonstrate that Twist1 deletion results in a significantly decreased long-term HSC (LT-HSC) frequency, markedly reduced dormancy and self-renewal capacities and skewed myeloid differentiation in steady-state hematopoiesis. Twist1-deficient LT-HSC are more compromised in tolerance of irradiation and 5 fluorouracil-induced stresses, and exhibit typical phenotypes of senescence and higher levels of DNA damage and apoptosis. Mechanistically, Twist1 deficiency upregulates the expression of voltage-gated calcium channel Cacna1b in HSC, leading to noticeable increases in mitochondrial calcium levels, biogenesis, metabolic activity and reactive oxygen species production. Suppression of voltage-gated calcium channel by a calcium channel blocker largely rescues the phenotypic and functional defects in Twist1-deleted HSCs under both steady-state and stress conditions. Collectively, our data, for the first time, characterize TWIST1 as a critical regulator of HSC function acting through CACNA1B/Ca2+/mitochondria axis, and highlight the importance of Ca2+ in HSC maintenance. These observations provide new insights into the mechanisms for the control of HSC fate. Disclosures No relevant conflicts of interest to declare.

Asymmetric organelle inheritance predicts human blood stem cell fate

Blood ◽  
2021 ◽  
Author(s):  
Dirk Loeffler ◽  
Florin Schneiter ◽  
Weijia Wang ◽  
Arne Wehling ◽  
Tobias Kull ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Cell Division ◽  
Cell Fate ◽  
Human Blood ◽  
Asymmetric Cell Division ◽  
Cell Fates ◽  
Hematopoietic Stem ◽  
Stem Cell Fate ◽  
Blood Stem Cells

Understanding human hematopoietic stem cell fate control is important for their improved therapeutic manipulation. Asymmetric cell division, the asymmetric inheritance of factors during division instructing future daughter cell fates, was recently described in mouse blood stem cells. In human blood stem cells, the possible existence of asymmetric cell division remained unclear due to technical challenges in its direct observation. Here, we use long-term quantitative single-cell imaging to show that lysosomes and active mitochondria are asymmetrically inherited in human blood stem cells and that their inheritance is a coordinated, non-random process. Furthermore, multiple additional organelles, including autophagosomes, mitophagosomes, autolysosomes and recycling endosomes show preferential asymmetric co-segregation with lysosomes. Importantly, asymmetric lysosomal inheritance predicts future asymmetric daughter cell cycle length, differentiation and stem cell marker expression, while asymmetric inheritance of active mitochondria correlates with daughter metabolic activity. Hence, human hematopoietic stem cell fates are regulated by asymmetric cell division, with both mechanistic evolutionary conservation and differences to the mouse system.

scholarly journals Human cytomegalovirus induces and exploits Roquin to counteract the IRF1-mediated antiviral state

2019 ◽  
Vol 116 (37) ◽  
pp. 18619-18628 ◽  
Author(s):  
Jaewon Song ◽  
Sanghyun Lee ◽  
Dong-Yeon Cho ◽  
Sungwon Lee ◽  
Hyewon Kim ◽  
...  
Keyword(s):  
Human Cytomegalovirus ◽  
Rna Processing ◽  
Rna Binding ◽  
Binding Protein ◽  
Rna Binding Protein ◽  
Transcriptome Profiling ◽  
Loss Of Function ◽  
Antiviral Genes ◽  
Cellular Antiviral Response ◽  
Infected Cells

RNA represents a pivotal component of host–pathogen interactions. Human cytomegalovirus (HCMV) infection causes extensive alteration in host RNA metabolism, but the functional relationship between the virus and cellular RNA processing remains largely unknown. Through loss-of-function screening, we show that HCMV requires multiple RNA-processing machineries for efficient viral lytic production. In particular, the cellular RNA-binding protein Roquin, whose expression is actively stimulated by HCMV, plays an essential role in inhibiting the innate immune response. Transcriptome profiling revealed Roquin-dependent global down-regulation of proinflammatory cytokines and antiviral genes in HCMV-infected cells. Furthermore, using cross-linking immunoprecipitation (CLIP)-sequencing (seq), we identified IFN regulatory factor 1 (IRF1), a master transcriptional activator of immune responses, as a Roquin target gene. Roquin reduces IRF1 expression by directly binding to its mRNA, thereby enabling suppression of a variety of antiviral genes. This study demonstrates how HCMV exploits host RNA-binding protein to prevent a cellular antiviral response and offers mechanistic insight into the potential development of CMV therapeutics.

scholarly journals Programmable RNA-binding protein composed of repeats of a single modular unit

2016 ◽  
Vol 113 (19) ◽  
pp. E2579-E2588 ◽  
Author(s):  
Katarzyna P. Adamala ◽  
Daniel A. Martin-Alarcon ◽  
Edward S. Boyden
Keyword(s):  
Rna Binding ◽  
Rna Binding Proteins ◽  
Binding Protein ◽  
Rna Binding Protein ◽  
Living Cells ◽  
Target Sequence ◽  
Rna Sequences ◽  
Intact Cells ◽  
Rna Translation ◽  
Protein Architecture

The ability to monitor and perturb RNAs in living cells would benefit greatly from a modular protein architecture that targets unmodified RNA sequences in a programmable way. We report that the RNA-binding protein PumHD (Pumilio homology domain), which has been widely used in native and modified form for targeting RNA, can be engineered to yield a set of four canonical protein modules, each of which targets one RNA base. These modules (which we call Pumby, for Pumilio-based assembly) can be concatenated in chains of varying composition and length, to bind desired target RNAs. The specificity of such Pumby–RNA interactions was high, with undetectable binding of a Pumby chain to RNA sequences that bear three or more mismatches from the target sequence. We validate that the Pumby architecture can perform RNA-directed protein assembly and enhancement of translation of RNAs. We further demonstrate a new use of such RNA-binding proteins, measurement of RNA translation in living cells. Pumby may prove useful for many applications in the measurement, manipulation, and biotechnological utilization of unmodified RNAs in intact cells and systems.

scholarly journals Genetic and Epigenetic Mechanisms That Maintain Hematopoietic Stem Cell Function

2016 ◽  
Vol 2016 ◽  
pp. 1-14 ◽  
Author(s):  
Christian Kosan ◽  
Maren Godmann
Keyword(s):  
Dna Damage ◽  
Stem Cell ◽  
Cell Fate ◽  
Molecular Mechanisms ◽  
Cell Function ◽  
Quiescent State ◽  
Ongoing Research ◽  
Hematopoietic Stem ◽  
Lineage Differentiation ◽  
Multipotent Progenitor Cells

All hematopoiesis cells develop from multipotent progenitor cells. Hematopoietic stem cells (HSC) have the ability to develop into all blood lineages but also maintain their stemness. Different molecular mechanisms have been identified that are crucial for regulating quiescence and self-renewal to maintain the stem cell pool and for inducing proliferation and lineage differentiation. The stem cell niche provides the microenvironment to keep HSC in a quiescent state. Furthermore, several transcription factors and epigenetic modifiers are involved in this process. These create modifications that regulate the cell fate in a more or less reversible and dynamic way and contribute to HSC homeostasis. In addition, HSC respond in a unique way to DNA damage. These mechanisms also contribute to the regulation of HSC function and are essential to ensure viability after DNA damage. How HSC maintain their quiescent stage during the entire life is still matter of ongoing research. Here we will focus on the molecular mechanisms that regulate HSC function.

GATA-2 Regulates Granulocyte-Macrophage Progenitor Cell Function.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1411-1411
Author(s):  
Neil P Rodrigues ◽  
Ashleigh S Boyd ◽  
Cristina Fugazza ◽  
Gillian E May ◽  
YanPing Guo ◽  
...  
Keyword(s):  
Cell Fate ◽  
Progenitor Cell ◽  
Cell Function ◽  
Hematopoietic Progenitor Cell ◽  
Cell Cycle Distribution ◽  
Functional Impairments ◽  
Hematopoietic Stem ◽  
Myeloid Progenitor ◽  
Functionally Impaired ◽  
Functional Defect

Abstract The zinc finger transcription factor GATA-2 has been implicated in the regulation of hematopoietic stem cells. Herein we explored the role of GATA-2 as a candidate regulator of the hematopoietic progenitor cell compartment. We showed that bone marrow from GATA-2 heterozygote (GATA-2+/-) mice displayed attenuated granulocyte-macrophage progenitor function in colony-forming cell (CFC) and serial replating CFC assays. This defect was mapped to the Lin−CD117+Sca-1−CD34+CD16/32high granulocyte-macrophage progenitor (GMP) compartment of GATA-2+/− marrow, which was reduced in size and functionally impaired in CFC assays and competitive transplantation. Similar functional impairments were obtained using a RNA interference approach to stably knockdown GATA-2 in wild-type GMP. While apoptosis and cell cycle distribution remained unperturbed in GATA-2+/− GMP, quiescent cells from GATA-2+/− GMP exhibited altered functionality. Gene expression analysis revealed attenuated expression of HES-1 mRNA in GATA-2 deficient GMPs. Binding of GATA-2 to the HES-1 locus was detected in the myeloid progenitor cell line 32Dcl3 and enforced expression of HES-1 expression in GATA-2+/− GMP rectified the functional defect, suggesting that GATA-2 regulates myeloid progenitor function through HES-1. These data collectively point to GATA-2 as novel, pivotal determinant of GMP cell fate.

A PML–PPAR-δ Pathway for Fatty Acid Oxidation Regulates Hematopoietic Stem Cell Maintenance Through the Control of Asymmetric Division.

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 2327-2327
Author(s):  
Keisuke Ito ◽  
Arkaitz Carracedo ◽  
Fumio Arai ◽  
Ugo Ala ◽  
David Avigan ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Research Funding ◽  
Asymmetric Cell Division ◽  
Cell Function ◽  
Asymmetric Division ◽  
Promyelocytic Leukemia ◽  
Acid Oxidation ◽  
Hematopoietic Stem ◽  
Daughter Cells

Abstract Abstract 2327 Stem-cell function is an exquisitely regulated process. Thus far, the contribution of metabolic cues to stem-cell function has not been well understood. Here we have established a new assay for the assessment of asymmetric division in hematopoietic stem cells (HSCs), and identify a previously unknown promyelocytic leukemia (PML)–peroxisome proliferator-activated receptor δ (PPAR-δ)–fatty-acid oxidation (FAO) pathway for the maintenance of HSCs and the control of asymmetric cell division. HSCs exist in a quiescent state in the bone marrow niche and are the source of all hematological progenitors and differentiated cells throughout the lifespan of an organism. One of the central tasks of stem-cell biology is to define the modes and mechanisms that regulate the self-renewal and commitment of stem cells, as alterations in this equilibrium have a substantial effect on hematopoietic homeostasis and maintenance. It has been suggested that asymmetric division of HSCs ensures that a fraction of daughter cells retain features of stem cells while replenishing the committed compartment of hematopoietic progenitors. Therefore, identifying the factors that regulate this process would be of great biological and therapeutic relevance. The PML tumor-suppressor gene, originally cloned at the break point of the t(15;17) chromosomal translocation of acute promyelocytic leukemia, has a key role in the maintenance of HSCs. However, how PML exerts its crucial function in the biology and maintenance of HSCs has not been established. In this study, we will present a new metabolic pathway downstream of PML, which can be regarded as a fine-tuning rheostat essential for HSC maintenance and their asymmetric division. Specifically, we find that loss of PPARδ or inhibition of mitochondrial FAO induces loss of HSC maintenance both in vitro and in vivo, whereas treatment with PPARδ agonists improved HSC maintenance. We demonstrate that PML exerts its essential role in HSC maintenance through regulation of PPAR signaling and FAO. Mechanistically, we show that the PML–PPAR δ–FAO pathway controls the asymmetric division of HSCs. Deletion of the Ppard or Pml genes as well as inhibition of FAO results in the symmetric commitment of HSC daughter cells, whereas PPARδ activation increases asymmetric cell division, and rescues the defect exhibited by Pml-deficient HSCs, therefore ensuring the correct maintenance of the HSC population. Thus, our findings identify a metabolic switch for the control of HSC cell fate with potential therapeutic implications. Disclosures: Avigan: Curetech: Research Funding; Celgene: Research Funding; Genzyme: Membership on an entity's Board of Directors or advisory committees.

Sign in / Sign up

Export Citation Format

Share Document