bims-scepro Biomed News
on Stem cell proteostasis
Issue of 2024–06–02
twenty-two papers selected by
William Grey, University of York
  1. Enhancing prime editing in hematopoietic stem and progenitor cells by modulating nucleotide metabolism.
  2. CRISPR Dependency Screens in Primary Hematopoietic Stem Cells Identify KDM3B as a Genotype Specific Vulnerability in IDH2- and TET2-Mutant Cells.
  3. BRD9 regulates normal human hematopoietic stem cell function and lineage differentiation.
  4. Chromatin modifier Hmga2 promotes adult hematopoietic stem cell function and blood regeneration in stress conditions.
  5. Temporal Single Cell Analysis of Leukemia Microenvironment Identifies Taurine-Taurine Transporter Axis as a Key Regulator of Myeloid Leukemia.
  6. Cell-type-specific consequences of mosaic structural variants in hematopoietic stem and progenitor cells.
  7. Adaptation to ex vivo culture reduces human hematopoietic stem cell activity independently of cell cycle.
  8. Tuning apicobasal polarity and junctional recycling in the hemogenic endothelium orchestrates the morphodynamic complexity of emerging pre-hematopoietic stem cells.
  9. LILRB4 represents a promising target for immunotherapy by dual targeting tumor cells and myeloid-derived suppressive cells in multiple myeloma.
  10. Unveiling mitochondrial and ribosomal gene deregulation and tumor microenvironment dynamics in acute myeloid leukemia.
  11. Alternative platelet differentiation pathways initiated by nonhierarchically related hematopoietic stem cells.
  12. The Hippo-YAP/TAZ-TEAD axis promotes multipotency in bone marrow stromal cells.
  13. Megakaryocytic IGF1 coordinates activation and ferroptosis to safeguard hematopoietic stem cell regeneration after radiation injury.
  14. Metabolic adaptation pilots the differentiation of human hematopoietic cells.
  15. Neutrophils cultured ex vivo from CD34+ stem cells are immature and genetically tractable.
  16. Asymmetric stem cell division maintains the genetic heterogeneity of tissue cells.
  17. EZH2 inhibition induces senescence via ERK1/2 signaling pathway in multiple myeloma.
  18. Inhibition of NOTCH4 sensitizes FLT3/ITD acute myeloid leukemia cells to FLT3 tyrosine kinase inhibition.
  19. In vivo Targeting MEK and TNK2/SRC pathways in PTPN11 driven leukemia.
  20. A novel function of STAT3β in suppressing interferon response improves outcome in acute myeloid leukemia.
  21. RUNX1::ETO and CBFβ::MYH11 converge on aberrant activation of BCAT1 to confer a therapeutic vulnerability in core-binding factor-acute myeloid leukaemia.
  22. Moesin activation controls bone resorption and tunneling nanotube-dependent osteoclast fusion.
  1. Nat Biotechnol. 2024 May 28.
    Enhancing prime editing in hematopoietic stem and progenitor cells by modulating nucleotide metabolism.
    Sébastien Levesque, Andrea Cosentino, Archana Verma, Pietro Genovese, Daniel E Bauer.
      Therapeutic prime editing of hematopoietic stem and progenitor cells (HSPCs) holds great potential to remedy blood disorders. Quiescent cells have low nucleotide levels and resist retroviral infection, and it is possible that nucleotide metabolism could limit reverse transcription-mediated prime editing in HSPCs. We demonstrate that deoxynucleoside supplementation and Vpx-mediated degradation of SAMHD1 improve prime editing efficiency in HSPCs, especially when coupled with editing approaches that evade mismatch repair.
    DOI:  https://doi.org/10.1038/s41587-024-02266-4
  2. Cancer Discov. 2024 May 31.
    CRISPR Dependency Screens in Primary Hematopoietic Stem Cells Identify KDM3B as a Genotype Specific Vulnerability in IDH2- and TET2-Mutant Cells.
    Michael R Waarts, Shoron Mowla, Meaghan Boileau, Anthony R Martinez Benitez, Junya Sango, Maya Bagish, Ines Fernandez-Maestre, Yufan Shan, Shira E Eisman, Young C Park, Matthew Wereski, Isabelle Csete, Kavi O'Connor, Angelica C Romero-Vega, Linde A Miles, Wenbin Xiao, Xiaodi Wu, Richard P Koche, Scott A Armstrong, Alan H Shih, Eirini P Papapetrou, Jason M Butler, Sheng F Cai, Robert L Bowman, Ross L Levine.
      Clonal hematopoiesis (CH) is a common premalignant state in the blood and confers an increased risk of blood cancers and all-cause mortality. Identification of therapeutic targets in CH has been hindered by the lack of an ex vivo platform amenable for studying primary hematopoietic stem and progenitor cells (HSPCs). Here, we utilize an ex vivo co-culture system of HSPCs with bone marrow endothelial cells to perform CRISPR/Cas9 screens in mutant HSPCs. Our data reveal that loss of the histone demethylase family members Kdm3b and Jmjd1c specifically reduces the fitness of Idh2- and Tet2-mutant HSPCs. Kdm3b loss in mutant cells leads to decreased expression of critical cytokine receptors including Mpl, rendering mutant HSPCs preferentially susceptible to inhibition of downstream JAK2 signaling. Our study nominates an epigenetic regulator and an epigenetically regulated receptor signaling pathway as genotype-specific therapeutic targets and provides a scalable platform to identify genetic dependencies in mutant HSPCs.
    DOI:  https://doi.org/10.1158/2159-8290.CD-23-1092
  3. Cell Death Differ. 2024 May 30.
    BRD9 regulates normal human hematopoietic stem cell function and lineage differentiation.
    Swati Garg, Wei Ni, Basudev Chowdhury, Ellen L Weisberg, Martin Sattler, James D Griffin.
      Bromodomain containing protein 9 (BRD9), a member of the non-canonical BRG1/BRM-associated factor (ncBAF) chromatin remodeling complex, has been implicated as a synthetic lethal target in AML but its function in normal human hematopoiesis is unknown. In hematopoietic stem and progenitor cells (HSPC) genomic or chemical inhibition of BRD9 led to a proliferative disadvantage and loss of stem cells in vitro. Human HSPCs with reduced BRD9 protein levels produced lower numbers of immature mixed multipotent GEMM colonies in semi-solid media. In lineage-promoting culture conditions, cells with reduced BRD9 levels failed to differentiate into the megakaryocytic lineage and showed delayed differentiation into erythroid cells but enhanced terminal myeloid differentiation. HSPCs with BRD9 knock down (KD) had reduced long-term multilineage engraftment in a xenotransplantation assay. An increased number of downregulated genes in RNAseq analysis after BRD9 KD coupled with a gain in chromatin accessibility at the promoters of several repressive transcription factors (TF) suggest that BRD9 functions in the maintenance of active transcription during HSC differentiation. In particular, the hematopoietic master regulator GATA1 was identified as one of the core TFs regulating the gene networks modulated by BRD9 loss in HSPCs. BRD9 inhibition reduced a GATA1-luciferase reporter signal, further suggesting a role for BRD9 in regulating GATA1 activity. BRD9 is therefore an additional example of epigenetic regulation of human hematopoiesis.
    DOI:  https://doi.org/10.1038/s41418-024-01306-5
  4. EMBO J. 2024 May 29.
    Chromatin modifier Hmga2 promotes adult hematopoietic stem cell function and blood regeneration in stress conditions.
    Sho Kubota, Yuqi Sun, Mariko Morii, Jie Bai, Takako Ideue, Mayumi Hirayama, Supannika Sorin, Eerdunduleng, Takako Yokomizo-Nakano, Motomi Osato, Ai Hamashima, Mihoko Iimori, Kimi Araki, Terumasa Umemoto, Goro Sashida.
      The molecular mechanisms governing the response of hematopoietic stem cells (HSCs) to stress insults remain poorly defined. Here, we investigated effects of conditional knock-out or overexpression of Hmga2 (High mobility group AT-hook 2), a transcriptional activator of stem cell genes in fetal HSCs. While Hmga2 overexpression did not affect adult hematopoiesis under homeostasis, it accelerated HSC expansion in response to injection with 5-fluorouracil (5-FU) or in vitro treatment with TNF-α. In contrast, HSC and megakaryocyte progenitor cell numbers were decreased in Hmga2 KO animals. Transcription of inflammatory genes was repressed in Hmga2-overexpressing mice injected with 5-FU, and Hmga2 bound to distinct regions and chromatin accessibility was decreased in HSCs upon stress. Mechanistically, we found that casein kinase 2 (CK2) phosphorylates the Hmga2 acidic domain, promoting its access and binding to chromatin, transcription of anti-inflammatory target genes, and the expansion of HSCs under stress conditions. Notably, the identified stress-regulated Hmga2 gene signature is activated in hematopoietic stem progenitor cells of human myelodysplastic syndrome patients. In sum, these results reveal a TNF-α/CK2/phospho-Hmga2 axis controlling adult stress hematopoiesis.
    Keywords:  5-Fluorouracil; Casein Kinase 2; Megakaryocyte; Self-renewal; TNF-α
    DOI:  https://doi.org/10.1038/s44318-024-00122-4
  5. bioRxiv. 2024 May 14. pii: 2024.05.11.593633. [Epub ahead of print]
    Temporal Single Cell Analysis of Leukemia Microenvironment Identifies Taurine-Taurine Transporter Axis as a Key Regulator of Myeloid Leukemia.
    Benjamin J Rodems, Sonali Sharma, Cameron D Baker, Christina M Kaszuba, Takashi Ito, Jane L Liesveld, Laura M Calvi, Michael W Becker, Craig T Jordan, John M Ashton, Jeevisha Bajaj.
      Signals from the microenvironment are known to be critical for development, sustaining adult stem cells, and for oncogenic progression. While candidate niche-driven signals that can promote cancer progression have been identified 1-6 , concerted efforts to comprehensively map microenvironmental ligands for cancer stem cell specific surface receptors have been lacking. Here, we use temporal single cell RNA-sequencing to identify molecular cues from the bone marrow stromal niche that engage leukemia stem cells (LSC) during oncogenic progression. We integrate these data with our RNA-seq analysis of human LSCs from distinct aggressive myeloid cancer subtypes and our CRISPR based in vivo LSC dependency map 7 to develop a temporal receptor-ligand interactome essential for disease progression. These analyses identify the taurine transporter (TauT)-taurine axis as a critical dependency of myeloid malignancies. We show that taurine production is restricted to the osteolineage population during cancer initiation and expansion. Inhibiting taurine synthesis in osteolineage cells impairs LSC growth and survival. Our experiments with the TauT genetic loss of function murine model indicate that its loss significantly impairs the progression of aggressive myeloid leukemias in vivo by downregulating glycolysis. Further, TauT inhibition using a small molecule strongly impairs the growth and survival of patient derived myeloid leukemia cells. Finally, we show that TauT inhibition can synergize with the clinically approved oxidative phosphorylation inhibitor venetoclax 8, 9 to block the growth of primary human leukemia cells. Given that aggressive myeloid leukemias continue to be refractory to current therapies and have poor prognosis, our work indicates targeting the taurine transporter may be of therapeutic significance. Collectively, our data establishes a temporal landscape of stromal signals during cancer progression and identifies taurine-taurine transporter signaling as an important new regulator of myeloid malignancies.
    DOI:  https://doi.org/10.1101/2024.05.11.593633
  6. Nat Genet. 2024 May 28.
    Cell-type-specific consequences of mosaic structural variants in hematopoietic stem and progenitor cells.
    Karen Grimes, Hyobin Jeong, Amanda Amoah, Nuo Xu, Julian Niemann, Benjamin Raeder, Patrick Hasenfeld, Catherine Stober, Tobias Rausch, Eva Benito, Johann-Christoph Jann, Daniel Nowak, Ramiz Emini, Markus Hoenicka, Andreas Liebold, Anthony Ho, Shimin Shuai, Hartmut Geiger, Ashley D Sanders, Jan O Korbel.
      The functional impact and cellular context of mosaic structural variants (mSVs) in normal tissues is understudied. Utilizing Strand-seq, we sequenced 1,133 single-cell genomes from 19 human donors of increasing age, and discovered the heterogeneous mSV landscapes of hematopoietic stem and progenitor cells. While mSVs are continuously acquired throughout life, expanded subclones in our cohort are confined to individuals >60. Cells already harboring mSVs are more likely to acquire additional somatic structural variants, including megabase-scale segmental aneuploidies. Capitalizing on comprehensive single-cell micrococcal nuclease digestion with sequencing reference data, we conducted high-resolution cell-typing for eight hematopoietic stem and progenitor cells. Clonally expanded mSVs disrupt normal cellular function by dysregulating diverse cellular pathways, and enriching for myeloid progenitors. Our findings underscore the contribution of mSVs to the cellular and molecular phenotypes associated with the aging hematopoietic system, and establish a foundation for deciphering the molecular links between mSVs, aging and disease susceptibility in normal tissues.
    DOI:  https://doi.org/10.1038/s41588-024-01754-2
  7. Blood. 2024 May 28. pii: blood.2023021426. [Epub ahead of print]
    Adaptation to ex vivo culture reduces human hematopoietic stem cell activity independently of cell cycle.
    Carys S Johnson, Matthew J Williams, Kendig Sham, Serena Belluschi, Wenjuan Ma, Xiaonan Wang, Winnie Wy Lau, Kerstin Kaufmann, Gabriela Krivdova, Emily Francesca Calderbank, Nicole Mende, Jessica McLeod, Giovanna Mantica, Juan Li, Charlotte Grey-Wilson, Michael Drakopoulos, Shaaezmeen Basheer, Shubhankar Sinha, Evangelia Diamanti, Christina Basford, Nicola K Wilson, Steven J Howe, John E Dick, Berthold Gottgens, Anthony R Green, Natalie Francis, Elisa Laurenti.
      Loss of long-term hematopoietic stem cell (LT-HSC) function ex vivo hampers the success of clinical protocols reliant on culture. However, the kinetics and mechanisms by which this occurs remain incompletely characterized. Here, through time-resolved scRNA-Seq, matched in vivo functional analysis and the use of a reversible in vitro system of early G1 arrest, we define the sequence of transcriptional and functional events occurring during the first ex vivo division of human LT-HSCs. We demonstrate that the sharpest loss of LT-HSC repopulation capacity happens early on, between 6 and 24 hours of culture, before LT-HSCs commit to cell cycle progression. During this time window, LT-HSCs adapt to the culture environment, limiting global variability in gene expression and transiently upregulating gene networks involved in signaling and stress responses. From 24 hours, LT-HSC progression past early G1 contributes to the establishment of differentiation programmes in culture. However, contrary to current assumptions, we demonstrate that loss of HSC function ex vivo is independent of cell cycle progression. Finally, we show that targeting LT-HSC adaptation to culture by inhibiting early activation of JAK/STAT signaling improves HSC long-term repopulating function ex vivo. Collectively, our study demonstrates that controlling early LT-HSC adaptation to ex vivo culture, for example via JAK inhibition, is of critical importance to improve HSC gene therapy and expansion protocols.
    DOI:  https://doi.org/10.1182/blood.2023021426
  8. Elife. 2024 May 29. pii: RP91429. [Epub ahead of print]12
    Tuning apicobasal polarity and junctional recycling in the hemogenic endothelium orchestrates the morphodynamic complexity of emerging pre-hematopoietic stem cells.
    Léa Torcq, Sara Majello, Catherine Vivier, Anne A Schmidt.
      Hematopoietic stem cells emerge in the embryo from an aortic-derived tissue called the hemogenic endothelium (HE). The HE appears to give birth to cells of different nature and fate but the molecular principles underlying this complexity are largely unknown. Here we show, in the zebrafish embryo, that two cell types emerge from the aortic floor with radically different morphodynamics. With the support of live imaging, we bring evidence suggesting that the mechanics underlying the two emergence types rely, or not, on apicobasal polarity establishment. While the first type is characterized by reinforcement of apicobasal polarity and maintenance of the apical/luminal membrane until release, the second type emerges via a dynamic process reminiscent of trans-endothelial migration. Interfering with Runx1 function suggests that the balance between the two emergence types depends on tuning apicobasal polarity at the level of the HE. In support of this and unexpectedly, we show that Pard3ba - one of the four Pard3 proteins expressed in the zebrafish - is sensitive to interference with Runx1 activity, in aortic endothelial cells. This supports the idea of a signaling cross talk controlling cell polarity and its associated features, between aortic and hemogenic cells. In addition, using new transgenic fish lines that express Junctional Adhesion Molecules and functional interference, we bring evidence for the essential role of ArhGEF11/PDZ-RhoGEF in controlling the HE-endothelial cell dynamic interface, including cell-cell intercalation, which is ultimately required for emergence completion. Overall, we highlight critical cellular and dynamic events of the endothelial-to-hematopoietic transition that support emergence complexity, with a potential impact on cell fate.
    Keywords:  JAMs; Pard3; apico-basal polarity; developmental biology; endothelial-to-hematopoietic transition; hemogenic endothelium; junctional adhesion molecules; regenerative medicine; stem cell; stem cells; zebra fish (danio rerio)
    DOI:  https://doi.org/10.7554/eLife.91429
  9. Haematologica. 2024 May 30.
    LILRB4 represents a promising target for immunotherapy by dual targeting tumor cells and myeloid-derived suppressive cells in multiple myeloma.
    Lixin Gong, Hao Sun, Lanting Liu, Xiyue Sun, Teng Fang, Zhen Yu, Weiwei Sui, Jingyu Xu, Tingyu Wang, Fangshuo Feng, Lei Lei, Wei Rui, Yuxuan Liu, Xueqiang Zhao, Gang An, Xin Lin, Lugui Qiu, Mu Hao.
      Multiple myeloma (MM) remains an incurable hematological malignancy. Despite tremendous advances in the treatment, about 10% of patients still have very poor outcomes with median overall survival less than 24 months. Our study aimed to underscore the critical mechanisms pertaining to the rapid disease progression and provide novel therapeutic selection for these ultra-high-risk patients. We utilized single-cell transcriptomic sequencing to dissect the characteristic bone marrow niche of patients with survival of less than two years (EM24). Notably, an enrichment of LILRB4high pre-matured plasma-cell cluster was observed in the patients in EM24 compared to patients with durable remission. This cluster exhibited aggressive proliferation and drug-resistance phenotype. High-level LILRB4 promoted MM clonogenicity and progression. Clinically, high expression of LILRB4 was correlated with poor prognosis in both newly diagnosed MM patients and relapsed/refractory MM patients. The ATAC-seq analysis identified that high chromosomal accessibility caused the elevation of LILRB4 on MM cells. CRISPR-Cas9 deletion of LILRB4 alleviated the growth of MM cells, inhibited the immunosuppressive function of MDSCs, and further rescued T cell dysfunction in MM microenvironment. The more infiltration of myeloid-derived suppressive cells (MDSCs) was observed in EM24 patients as well. Therefore, we innovatively generated a TCR-based chimeric antigen receptor (CAR) T cell, LILRB4-STAR-T. Cytotoxicity experiment demonstrated that LILRB4-STAR-T cells efficaciously eliminated tumor cells and impeded MDSCs function. In conclusion, our study elucidates that LILRB4 is an ideal biomarker and promising immunotherapy target for high-risk MM. LILRB4-STAR-T cell immunotherapy is promising against tumor cells and immunosuppressive tumor microenvironment in MM.
    DOI:  https://doi.org/10.3324/haematol.2024.285099
  10. Cancer Gene Ther. 2024 May 28.
    Unveiling mitochondrial and ribosomal gene deregulation and tumor microenvironment dynamics in acute myeloid leukemia.
    Chao Ma, Yuchao Hao, Bo Shi, Zheng Wu, Di Jin, Xiao Yu, Bilian Jin.
      Acute myeloid leukemia (AML) is a malignant clonal hematopoietic disease with a poor prognosis. Understanding the interaction between leukemic cells and the tumor microenvironment (TME) can help predict the prognosis of leukemia and guide its treatment. Re-analyzing the scRNA-seq data from the CSC and G20 cohorts, using a Python-based pipeline including machine-learning-based scVI-tools, recapitulated the distinct hierarchical structure within the samples of AML patients. Weighted correlation network analysis (WGCNA) was conducted to construct a weighted gene co-expression network and to identify gene modules primarily focusing on hematopoietic stem cells (HSCs), multipotent progenitors (MPPs), and natural killer (NK) cells. The analysis revealed significant deregulation in gene modules associated with aerobic respiration and ribosomal/cytoplasmic translation. Cell-cell communications were elucidated by the CellChat package, revealing an imbalance of activating and inhibitory immune signaling pathways. Interception of genes upregulated in leukemic HSCs & MPPs as well as in NKG2A-high NK cells was used to construct prognostic models. Normal Cox and artificial neural network models based on 10 genes were developed. The study reveals the deregulation of mitochondrial and ribosomal genes in AML patients and suggests the co-occurrence of stimulatory and inhibitory factors in the AML TME.
    DOI:  https://doi.org/10.1038/s41417-024-00788-2
  11. Nat Immunol. 2024 May 30.
    Alternative platelet differentiation pathways initiated by nonhierarchically related hematopoietic stem cells.
    Joana Carrelha, Stefania Mazzi, Axel Winroth, Michael Hagemann-Jensen, Christoph Ziegenhain, Kari Högstrand, Masafumi Seki, Margs S Brennan, Madeleine Lehander, Bishan Wu, Yiran Meng, Ellen Markljung, Ruggiero Norfo, Hisashi Ishida, Karin Belander Strålin, Francesca Grasso, Christina Simoglou Karali, Affaf Aliouat, Amy Hillen, Edwin Chari, Kimberly Siletti, Supat Thongjuea, Adam J Mead, Sten Linnarsson, Claus Nerlov, Rickard Sandberg, Tetsuichi Yoshizato, Petter S Woll, Sten Eirik W Jacobsen.
      Rare multipotent stem cells replenish millions of blood cells per second through a time-consuming process, passing through multiple stages of increasingly lineage-restricted progenitors. Although insults to the blood-forming system highlight the need for more rapid blood replenishment from stem cells, established models of hematopoiesis implicate only one mandatory differentiation pathway for each blood cell lineage. Here, we establish a nonhierarchical relationship between distinct stem cells that replenish all blood cell lineages and stem cells that replenish almost exclusively platelets, a lineage essential for hemostasis and with important roles in both the innate and adaptive immune systems. These distinct stem cells use cellularly, molecularly and functionally separate pathways for the replenishment of molecularly distinct megakaryocyte-restricted progenitors: a slower steady-state multipotent pathway and a fast-track emergency-activated platelet-restricted pathway. These findings provide a framework for enhancing platelet replenishment in settings in which slow recovery of platelets remains a major clinical challenge.
    DOI:  https://doi.org/10.1038/s41590-024-01845-6
  12. bioRxiv. 2024 May 17. pii: 2024.05.14.594149. [Epub ahead of print]
    The Hippo-YAP/TAZ-TEAD axis promotes multipotency in bone marrow stromal cells.
    Yu-Hao Cheng, Chun Hei Ryan Chan, Qin Bian, Randall K Merling, Sergei A Kuznetsov, Pamela G Robey, Patrick Cahan.
      The mechanisms by which bone marrow stromal cells (BMSCs) maintain multilineage potency in vitro remain elusive. To identify the transcriptional regulatory circuits that contribute to BMSC multipotency, we performed paired single-nucleus multiomics of the expansion of freshly isolated BMSCs and of BMSCs undergoing tri-lineage differentiation. By computationally reconstructing the regulatory programs associated with initial stages of differentiation and early expansion, we identified the TEAD family of transcription factors, which is inhibited by Hippo signaling, as highly active in the BMSC in vitro multipotent state. Pharmacological inhibition of TEAD enhanced BMSC osteogenic and adipogenic differentiation, whereas its activation maintained BMSCs in an undifferentiated state, supporting a model whereby isolation of BMSCs coincides with a TEAD-controlled transcriptional state linked to multipotency. Our study highlights the Hippo pathway as a pivotal regulator of BMSC multipotency, and our regulatory network inferences are a reservoir of testable hypotheses that link transcription factors and their regulons to specific aspects of BMSC behavior.
    DOI:  https://doi.org/10.1101/2024.05.14.594149
  13. Cell Commun Signal. 2024 May 27. 22(1): 292
    Megakaryocytic IGF1 coordinates activation and ferroptosis to safeguard hematopoietic stem cell regeneration after radiation injury.
    Weinian Liao, Xinliang Chen, Shuzhen Zhang, Jun Chen, Chaonan Liu, Kuan Yu, Yimin Zhang, Mo Chen, Fang Chen, Mingqiang Shen, Binghui Lu, Songling Han, Song Wang, Junping Wang, Changhong Du.
       BACKGROUND: Hematopoietic stem cell (HSC) regeneration underlies hematopoietic recovery from myelosuppression, which is a life-threatening side effect of cytotoxicity. HSC niche is profoundly disrupted after myelosuppressive injury, while if and how the niche is reshaped and regulates HSC regeneration are poorly understood.
    METHODS: A mouse model of radiation injury-induced myelosuppression was built by exposing mice to a sublethal dose of ionizing radiation. The dynamic changes in the number, distribution and functionality of HSCs and megakaryocytes were determined by flow cytometry, immunofluorescence, colony assay and bone marrow transplantation, in combination with transcriptomic analysis. The communication between HSCs and megakaryocytes was determined using a coculture system and adoptive transfer. The signaling mechanism was investigated both in vivo and in vitro, and was consolidated using megakaryocyte-specific knockout mice and transgenic mice.
    RESULTS: Megakaryocytes become a predominant component of HSC niche and localize closer to HSCs after radiation injury. Meanwhile, transient insulin-like growth factor 1 (IGF1) hypersecretion is predominantly provoked in megakaryocytes after radiation injury, whereas HSCs regenerate paralleling megakaryocytic IGF1 hypersecretion. Mechanistically, HSCs are particularly susceptible to megakaryocytic IGF1 hypersecretion, and mTOR downstream of IGF1 signaling not only promotes activation including proliferation and mitochondrial oxidative metabolism of HSCs, but also inhibits ferritinophagy to restrict HSC ferroptosis. Consequently, the delicate coordination between proliferation, mitochondrial oxidative metabolism and ferroptosis ensures functional HSC expansion after radiation injury. Importantly, punctual IGF1 administration simultaneously promotes HSC regeneration and hematopoietic recovery after radiation injury, representing a superior therapeutic approach for myelosuppression.
    CONCLUSIONS: Our study identifies megakaryocytes as a last line of defense against myelosuppressive injury and megakaryocytic IGF1 as a novel niche signal safeguarding HSC regeneration.
    Keywords:  Ferroptosis; Hematopoietic stem cell; IGF1; Ionizing radiation; Megakaryocyte; Myelosuppression
    DOI:  https://doi.org/10.1186/s12964-024-01651-5
  14. Life Sci Alliance. 2024 Aug;pii: e202402747. [Epub ahead of print]7(8):
    Metabolic adaptation pilots the differentiation of human hematopoietic cells.
    Laëtitia Racine, Romuald Parmentier, Shreyas Niphadkar, Julie Chhun, Jean-Alain Martignoles, François Delhommeau, Sunil Laxman, Andras Paldi.
      A continuous supply of energy is an essential prerequisite for survival and represents the highest priority for the cell. We hypothesize that cell differentiation is a process of optimization of energy flow in a changing environment through phenotypic adaptation. The mechanistic basis of this hypothesis is provided by the established link between core energy metabolism and epigenetic covalent modifications of chromatin. This theory predicts that early metabolic perturbations impact subsequent differentiation. To test this, we induced transient metabolic perturbations in undifferentiated human hematopoietic cells using pharmacological inhibitors targeting key metabolic reactions. We recorded changes in chromatin structure and gene expression, as well as phenotypic alterations by single-cell ATAC and RNA sequencing, time-lapse microscopy, and flow cytometry. Our observations suggest that these metabolic perturbations are shortly followed by alterations in chromatin structure, leading to changes in gene expression. We also show that these transient fluctuations alter the differentiation potential of the cells.
    DOI:  https://doi.org/10.26508/lsa.202402747
  15. J Transl Med. 2024 May 31. 22(1): 526
    Neutrophils cultured ex vivo from CD34+ stem cells are immature and genetically tractable.
    Claire A Naveh, Kiran Roberts, Przemysław Zakrzewski, Christopher M Rice, Fernando M Ponce-Garcia, Kathryn Fleming, Megan Thompson, Nawamin Panyapiean, Huan Jiang, Stephanie Diezmann, Pedro L Moura, Ashley M Toye, Borko Amulic.
       BACKGROUND: Neutrophils are granulocytes with essential antimicrobial effector functions and short lifespans. During infection or sterile inflammation, emergency granulopoiesis leads to release of immature neutrophils from the bone marrow, serving to boost circulating neutrophil counts. Steady state and emergency granulopoiesis are incompletely understood, partly due to a lack of genetically amenable models of neutrophil development.
    METHODS: We optimised a method for ex vivo production of human neutrophils from CD34+ haematopoietic progenitors. Using flow cytometry, we phenotypically compared cultured neutrophils with native neutrophils from donors experiencing emergency granulopoiesis, and steady state neutrophils from non-challenged donors. We carry out functional and proteomic characterisation of cultured neutrophils and establish genome editing of progenitors.
    RESULTS: We obtain high yields of ex vivo cultured neutrophils, which phenotypically resemble immature neutrophils released into the circulation during emergency granulopoiesis. Cultured neutrophils have similar rates of ROS production and bacterial killing but altered degranulation, cytokine release and antifungal activity compared to mature neutrophils isolated from peripheral blood. These differences are likely due to incomplete synthesis of granule proteins, as demonstrated by proteomic analysis.
    CONCLUSION: Ex vivo cultured neutrophils are genetically tractable via genome editing of precursors and provide a powerful model system for investigating the properties and behaviour of immature neutrophils.
    Keywords:  CD34+stem cells; Ex vivo differentiation; Granulopoiesis; Neutrophil; Proteomics
    DOI:  https://doi.org/10.1186/s12967-024-05337-x
  16. bioRxiv. 2024 May 16. pii: 2024.05.16.594576. [Epub ahead of print]
    Asymmetric stem cell division maintains the genetic heterogeneity of tissue cells.
    Muhammed Burak Bener, Boris M Slepchenko, Mayu Inaba.
      Within a given tissue, the stem cell niche provides the microenvironment for stem cells suitable for their self-renewal. Conceptually, the niche space constrains the size of a stem-cell pool, as the cells sharing the niche compete for its space. It has been suggested that either neutral- or non-neutral-competition of stem cells changes the clone dynamics of stem cells. Theoretically, if the rate of asymmetric division is high, the stem cell competition is limited, thus suppressing clonal expansion. However, the effects of asymmetric division on clone dynamics have never been experimentally tested. Here, using the Drosophila germline stem cell (GSC) system, as a simple model of the in-vivo niche, we examine the effect of division modes (asymmetric or symmetric) on clonal dynamics by combining experimental approaches with mathematical modeling. Our experimental data and computational model both suggest that the rate of asymmetric division is proportional to the time a stem cell clone takes to expand. Taken together, our data suggests that asymmetric division is essential for maintaining the genetic variation of stem cells and thus serves as a critical mechanism for safeguarding fertility over the animal age or preventing multiple disorders caused by the clonal expansion of stem cells.
    DOI:  https://doi.org/10.1101/2024.05.16.594576
  17. Acta Biochim Biophys Sin (Shanghai). 2024 May 27.
    EZH2 inhibition induces senescence via ERK1/2 signaling pathway in multiple myeloma.
    Shushan Guo, Qiongwei Tang, Xuejie Gao, Liangning Hu, Ke Hu, Hui Zhang, Qikai Zhang, Yue Lai, Yujie Liu, Zhuning Wang, Shuaikang Chang, Yifei Zhang, Huifang Hu, Dong An, Yu Peng, Haiyan Cai, Jumei Shi.
      Epigenetic modifications play an important role in cellular senescence, and enhancer of zeste homolog 2 (EZH2) is a key methyltransferase involved in epigenetic remodeling in multiple myeloma (MM) cells. We have previously demonstrated that GSK126, a specific EZH2 inhibitor, exhibits anti-MM therapeutic efficacy and safety in vivo and in vitro; however, its specific mechanism remains unclear. This study shows that GSK126 induces cellular senescence in MM, which is characterized by the accumulation of senescence-associated heterochromatin foci (SAHF) and p21, and increased senescence-associated β galactosidase activity. Furthermore, EZH2 is inhibited in ribonucleotide reductase regulatory subunit M2 (RRM2) overexpression OCI-MY5 and RPMI-8226 cells. RRM2 overexpression inhibits the methyltransferase function of EZH2 and promotes its degradation through the ubiquitin-proteasome pathway, thereby inducing cellular senescence. In this senescence model, Lamin B1, a key component of the nuclear envelope and a marker of senescence, does not decrease but instead undergoes aberrant accumulation. Meanwhile, phosphorylation of extracellular signal-regulated protein kinase (ERK1/2) is significantly increased. The inhibition of ERK1/2 phosphorylation in turn partially restores Lamin B1 level and alleviates senescence. These findings suggest that EZH2 inhibition increases Lamin B1 level and induces senescence by promoting ERK1/2 phosphorylation. These data indicate that EZH2 plays an important role in MM cellular senescence and provide insights into the relationships among Lamin B1, p-ERK1/2, and cellular senescence.
    Keywords:  EZH2; Lamin B1; RRM2; cellular senescence; multiple myeloma
    DOI:  https://doi.org/10.3724/abbs.2024077
  18. Leukemia. 2024 May 29.
    Inhibition of NOTCH4 sensitizes FLT3/ITD acute myeloid leukemia cells to FLT3 tyrosine kinase inhibition.
    Ruiqi Zhu, Courtney M Shirley, S Haihua Chu, Li Li, Bao H Nguyen, Jaesung Seo, Min Wu, Tessa Seale, Amy S Duffield, Louis M Staudt, Mark Levis, Yu Hu, Donald Small.
      Internal tandem duplication mutations of FLT3 (FLT3/ITD) confer poor prognosis in AML. FLT3 tyrosine kinase inhibitors (TKIs) alone have limited and transient clinical efficacy thus calling for new targets for more effective combination therapy. In a loss-of-function RNAi screen, we identified NOTCH4 as one such potential target whose inhibition proved cytotoxic to AML cells, and also sensitized them to FLT3 inhibition. Further investigation found increased NOTCH4 expression in FLT3/ITD AML cell lines and primary patient samples. Inhibition of NOTCH4 by shRNA knockdown, CRISPR-Cas9-based knockout or γ-secretase inhibitors synergized with FLT3 TKIs to kill FLT3/ITD AML cells in vitro. NOTCH4 inhibition sensitized TKI-resistant FLT3/ITD cells to FLT3 TKI inhibition. The combination reduced phospho-ERK and phospho-AKT, indicating inhibition of MAPK and PI3K/AKT signaling pathways. It also led to changes in expression of genes involved in regulating cell cycling, DNA repair and transcription. A patient-derived xenograft model showed that the combination reduced both the level of leukemic involvement of primary human FLT3/ITD AML cells and their ability to engraft secondary recipients. In summary, these results demonstrate that NOTCH4 inhibition synergizes with FLT3 TKIs to eliminate FLT3/ITD AML cells, providing a new therapeutic target for AML with FLT3/ITD mutations.
    DOI:  https://doi.org/10.1038/s41375-024-02292-w
  19. bioRxiv. 2024 May 18. pii: 2024.05.16.594555. [Epub ahead of print]
    In vivo Targeting MEK and TNK2/SRC pathways in PTPN11 driven leukemia.
    Bill H Chang, Karina Thiel-Klare, Jeffrey W Tyner.
      PTPN11 encodes for a tyrosine phosphatase implicated in the pathogenesis of hematologic malignancies such as Juvenile Myelomonocytic Leukemia (JMML), Acute Myeloid Leukemia (AML), and Acute Lymphoblastic Leukemia (ALL). Since activating mutations of PTPN11 increase proliferative signaling and cell survival through the RAS/MAPK proliferative pathway there is significant interest in using MEK inhibitors for clinical benefit. Yet, single agent clinical activity has been minimal. Previously, we showed that PTPN11 is further activated by upstream tyrosine kinases TNK2/SRC, and that PTPN11-mutant JMML and AML cells are sensitive to TNK2 inhibition using dasatinib. In these studies, we adopted a genetically engineered mouse model of PTPN11 driven leukemia using the mouse strain 129S/Sv- Ptpn11 tm6Bgn /Mmucd crossed with B6.129P2- Lyz2 tm1(cre)Ifo /J. The F1 progeny expressing Ptpn11 D61Y within hematopoietic cells destined along the granulocyte-monocyte progenitor lineage developed a myeloproliferative disorder. Cohorts of animals treated with combination of dasatinib and trametinib had a significant effect in mitigating disease parameters compared to single agents. Further, a primary patient-derived xenograft model using a myeloid leukemia with PTPN11 F71L also displayed improved response to combination. Collectively, these studies point to combined therapies targeting MEK and TNK2/SRC as a promising therapeutic potential for PTPN11-mutant leukemias.
    Key Points: Combining MEK and TNK2/SRC inhibitors has therapeutic potential in PTPN11 mutant JMML and AML.
    DOI:  https://doi.org/10.1101/2024.05.16.594555
  20. Cell Death Dis. 2024 May 28. 15(5): 369
    A novel function of STAT3β in suppressing interferon response improves outcome in acute myeloid leukemia.
    Sophie Edtmayer, Agnieszka Witalisz-Siepracka, Bernhard Zdársky, Kerstin Heindl, Stefanie Weiss, Thomas Eder, Sayantanee Dutta, Uwe Graichen, Sascha Klee, Omar Sharif, Rotraud Wieser, Balázs Győrffy, Valeria Poli, Emilio Casanova, Heinz Sill, Florian Grebien, Dagmar Stoiber.
      Signal transducer and activator of transcription 3 (STAT3) is frequently overexpressed in patients with acute myeloid leukemia (AML). STAT3 exists in two distinct alternatively spliced isoforms, the full-length isoform STAT3α and the C-terminally truncated isoform STAT3β. While STAT3α is predominantly described as an oncogenic driver, STAT3β has been suggested to act as a tumor suppressor. To elucidate the role of STAT3β in AML, we established a mouse model of STAT3β-deficient, MLL-AF9-driven AML. STAT3β deficiency significantly shortened survival of leukemic mice confirming its role as a tumor suppressor. Furthermore, RNA sequencing revealed enhanced STAT1 expression and interferon (IFN) signaling upon loss of STAT3β. Accordingly, STAT3β-deficient leukemia cells displayed enhanced sensitivity to blockade of IFN signaling through both an IFNAR1 blocking antibody and the JAK1/2 inhibitor Ruxolitinib. Analysis of human AML patient samples confirmed that elevated expression of IFN-inducible genes correlated with poor overall survival and low STAT3β expression. Together, our data corroborate the tumor suppressive role of STAT3β in a mouse model in vivo. Moreover, they provide evidence that its tumor suppressive function is linked to repression of the STAT1-mediated IFN response. These findings suggest that the STAT3β/α mRNA ratio is a significant prognostic marker in AML and holds crucial information for targeted treatment approaches. Patients displaying a low STAT3β/α mRNA ratio and unfavorable prognosis could benefit from therapeutic interventions directed at STAT1/IFN signaling.
    DOI:  https://doi.org/10.1038/s41419-024-06749-9
  21. Br J Haematol. 2024 May 27.
    RUNX1::ETO and CBFβ::MYH11 converge on aberrant activation of BCAT1 to confer a therapeutic vulnerability in core-binding factor-acute myeloid leukaemia.
    Siyang Wang, Yabin Liu, Xujie Zhao, Xiaoling Wang, Jiacheng Lou, Peng Jin, Yi Zhang, Jinyi Yu, Kankan Wang.
      Effectively targeting transcription factors in therapeutic interventions remains challenging, especially in core-binding factor-acute myeloid leukaemia (CBF-AML) characterized by RUNX1::ETO and CBFβ::MYH11 fusions. However, recent studies have drawn attention towards aberrant amino acid metabolisms as actionable therapeutic targets. Here, by integrating the expression profile and genetic makeup in AML cohort, we found higher BCAT1 expression in CBF-AML patients compared with other subtypes. Metabolic profiling revealed that high BCAT1 expression led to reprogrammed branch amino acid metabolism in CBF-AML and was associated with sphingolipid pathway relating to the fitness of leukaemia cells, supported by transcriptomic profiling. Mechanistically, we demonstrated in cell lines and primary patient samples that BCAT1 was directly activated by RUNX1::ETO and CBFβ::MYH11 fusion proteins similarly in a RUNX1-dependent manner through rewiring chromatin conformation at the BCAT1 gene locus. Furthermore, BCAT1 inhibition resulted in blunted cell cycle, enhanced apoptosis and myeloid differentiation of CBF-AML cells in vitro, and alleviated leukaemia burden and prolonged survival in vivo. Importantly, pharmacological inhibition of BCAT1 using the specific inhibitor Gabapentin demonstrated therapeutic effects, as evidenced by delayed leukaemia progression and improved survival in vivo. In conclusion, our study uncovers BCAT1 as a genetic vulnerability and a promising targeted therapeutic opportunity for CBF-AML.
    Keywords:  BCAT1; CBFβ::MYH11; RUNX1::ETO; core‐binding factor‐acute myeloid leukaemia
    DOI:  https://doi.org/10.1111/bjh.19565
  22. bioRxiv. 2024 May 15. pii: 2024.05.13.593799. [Epub ahead of print]
    Moesin activation controls bone resorption and tunneling nanotube-dependent osteoclast fusion.
    Ophélie Dufrancais, Perrine Verdys, Arnaud Métais, Marie Juzans, Thibaut Sanchez, Martin Bergert, Marianna Plozza, Julia Halper, Christopher J Panebianco, Rémi Mascarau, Rémi Gence, Gaëlle Arnaud, Myriam Ben Neji, Isabelle Maridonneau-Parini, Véronique Le Cabec, Joel D Boerckel, Nathan J Pavlos, Alba Diz-Muñoz, Frédéric Lagarrigue, Claudine Blin-Wakkach, Sébastien Carréno, Renaud Poincloux, Janis K Burkhardt, Brigitte Raynaud-Messina, Christel Vérollet.
      Osteoclasts are multinucleated cells unique in their ability to resorb bone. Osteoclastogenesis involves several steps of actin-driven rearrangements that participate not only in the cell-cell fusion process, but also in the formation of the sealing zone, the adhesive structure determining the resorption area. Despite the importance of these actin cytoskeleton-based processes, their precise mechanisms of regulation are still poorly characterized. Here, we found that moesin, a member of the Ezrin/Radixin/Moesin (ERM) protein family, is activated during osteoclast maturation and plays an instrumental role for both osteoclast fusion and function. In mouse and human osteoclast precursors, moesin is negatively regulated to potentiate their ability to fuse and degrade bone. Accordingly, we demonstrated that moesin depletion decreases membrane-to-cortex attachment and enhances formation of tunneling nanotubes (TNTs), F-actin-containing intercellular bridges that we revealed to trigger osteoclast fusion. In addition, via a β3-integrin/RhoA/SLK pathway and independently of its role in fusion, moesin regulates the number and organization of sealing zones in mature osteoclast, and thus participates in the control of bone resorption. Supporting these findings, we found that moesin-deficient mice are osteopenic with a reduced density of trabecular bones and increased osteoclast abundance and activity. These findings provide a better understanding of the regulation of osteoclast biology, and open new opportunities to specifically target osteoclast activity in bone disease therapy.
    DOI:  https://doi.org/10.1101/2024.05.13.593799
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