bims-scepro Biomed News
on Stem cell proteostasis
Issue of 2024–05–19
twenty papers selected by
William Grey, University of York
  1. Autophagy in hematopoietic stem cell development of the embryo.
  2. OGT and OGA gene-edited human induced pluripotent stem cells for dissecting the functional roles of O-GlcNAcylation in hematopoiesis.
  3. Autophagy counters inflammation-driven glycolytic impairment in aging hematopoietic stem cells.
  4. Age-specific induction of mutant p53 drives clonal hematopoiesis and acute myeloid leukemia in adult mice.
  5. Complexities of modelling the bone marrow microenvironment to facilitate haematopoietic research.
  6. Metabolic dependencies of acute myeloid leukemia stem cells.
  7. Progresses in overcoming the limitations of in vitro erythropoiesis using human induced pluripotent stem cells.
  8. PI3K/HSCB axis facilitates FOG1 nuclear translocation to promote erythropoiesis and megakaryopoiesis.
  9. PROTAC-induced Protein Functional Dynamics in Targeted Protein Degradation.
  10. Bone marrow stromal cells induce chromatin remodeling in multiple myeloma cells leading to transcriptional changes.
  11. IRE1α determines ferroptosis sensitivity through regulation of glutathione synthesis.
  12. Extracellular signals induce dynamic ER remodeling through αTAT1-dependent microtubule acetylation.
  13. Generation of allogeneic CAR-NKT cells from hematopoietic stem and progenitor cells using a clinically guided culture method.
  14. Deep PIM kinase substrate profiling reveals new rational co-therapeutic strategies for acute myeloid leukemia.
  15. High-throughput mechanical phenotyping and transcriptomics of single cells.
  16. Long-term hematopoietic transfer of the anti-cancer and lifespan-extending capabilities of a genetically engineered blood system by transplantation of bone marrow mononuclear cells.
  17. CD37 is a safe chimeric antigen receptor target to treat acute myeloid leukemia.
  18. A Guide to the Quantitation of Protein Secretion Dynamics at the Single-Cell Level.
  19. TIF1β activates leukemic transcriptional program in HSCs and promotes BCR::ABL1-induced myeloid leukemia.
  20. An age-progressive platelet differentiation path from hematopoietic stem cells causes exacerbated thrombosis.
  1. Autophagy. 2024 May 14.
    Autophagy in hematopoietic stem cell development of the embryo.
    Yumin Liu, Sifan Luo, Yuehang Chen, Zhuan Li.
      Hematopoietic stem cells (HSC) emerge from hemogenic endothelial cells (HEC) in the aorta-gonad-mesonephros (AGM) region of embryos, which go through the pre-HSC process. Various intrinsic and extrinsic factors are involved in this process. We recently discovered that the existence of distinct macroautophagic/autophagic statuses in hematopoietic precursors is related to the hematopoietic potential of pre-HSCs and the depletion of the Atg5 (autophagy related 5) gene specifically in endothelial cells impaired in the transition of endothelial to pre-HSCs, by hampering the autophagic process, likely via the NCL (nucleolin) pathway.
    Keywords:  Atg5; NCL; autophagy; hematopoietic development; hematopoietic precursors; hematopoietic stem cells
    DOI:  https://doi.org/10.1080/15548627.2024.2355072
  2. Front Cell Dev Biol. 2024 ;12 1361943
    OGT and OGA gene-edited human induced pluripotent stem cells for dissecting the functional roles of O-GlcNAcylation in hematopoiesis.
    Sudjit Luanpitpong, Kantpitchar Tangkiettrakul, Xing Kang, Pimonwan Srisook, Jirarat Poohadsuan, Parinya Samart, Phatchanat Klaihmon, Montira Janan, Chanchao Lorthongpanich, Chuti Laowtammathron, Surapol Issaragrisil.
      Hematopoiesis continues throughout life to produce all types of blood cells from hematopoietic stem cells (HSCs). Metabolic state is a known regulator of HSC self-renewal and differentiation, but whether and how metabolic sensor O-GlcNAcylation, which can be modulated via an inhibition of its cycling enzymes O-GlcNAcase (OGA) and O-GlcNAc transferase (OGT), contributes to hematopoiesis remains largely unknown. Herein, isogenic, single-cell clones of OGA-depleted (OGAi) and OGT-depleted (OGTi) human induced pluripotent stem cells (hiPSCs) were successfully generated from the master hiPSC line MUSIi012-A, which were reprogrammed from CD34+ hematopoietic stem/progenitor cells (HSPCs) containing epigenetic memory. The established OGAi and OGTi hiPSCs exhibiting an increase or decrease in cellular O-GlcNAcylation concomitant with their loss of OGA and OGT, respectively, appeared normal in phenotype and karyotype, and retained pluripotency, although they may favor differentiation toward certain germ lineages. Upon hematopoietic differentiation through mesoderm induction and endothelial-to-hematopoietic transition, we found that OGA inhibition accelerates hiPSC commitment toward HSPCs and that disruption of O-GlcNAc homeostasis affects their commitment toward erythroid lineage. The differentiated HSPCs from all groups were capable of giving rise to all hematopoietic progenitors, thus confirming their functional characteristics. Altogether, the established single-cell clones of OGTi and OGAi hiPSCs represent a valuable platform for further dissecting the roles of O-GlcNAcylation in blood cell development at various stages and lineages of blood cells. The incomplete knockout of OGA and OGT in these hiPSCs makes them susceptible to additional manipulation, i.e., by small molecules, allowing the molecular dynamics studies of O-GlcNAcylation.
    Keywords:  HSC; O-GlcNAcylation; OGA; OGT; differentiation; hematopoietic stem cell; hiPSC; induced pluripotent stem cell
    DOI:  https://doi.org/10.3389/fcell.2024.1361943
  3. Cell Stem Cell. 2024 May 13. pii: S1934-5909(24)00174-7. [Epub ahead of print]
    Autophagy counters inflammation-driven glycolytic impairment in aging hematopoietic stem cells.
    Paul V Dellorusso, Melissa A Proven, Fernando J Calero-Nieto, Xiaonan Wang, Carl A Mitchell, Felix Hartmann, Meelad Amouzgar, Patricia Favaro, Andrew DeVilbiss, James W Swann, Theodore T Ho, Zhiyu Zhao, Sean C Bendall, Sean Morrison, Berthold Göttgens, Emmanuelle Passegué.
      Autophagy is central to the benefits of longevity signaling programs and to hematopoietic stem cell (HSC) response to nutrient stress. With age, a subset of HSCs increases autophagy flux and preserves regenerative capacity, but the signals triggering autophagy and maintaining the functionality of autophagy-activated old HSCs (oHSCs) remain unknown. Here, we demonstrate that autophagy is an adaptive cytoprotective response to chronic inflammation in the aging murine bone marrow (BM) niche. We find that inflammation impairs glucose uptake and suppresses glycolysis in oHSCs through Socs3-mediated inhibition of AKT/FoxO-dependent signaling, with inflammation-mediated autophagy engagement preserving functional quiescence by enabling metabolic adaptation to glycolytic impairment. Moreover, we show that transient autophagy induction via a short-term fasting/refeeding paradigm normalizes glycolytic flux and significantly boosts oHSC regenerative potential. Our results identify inflammation-driven glucose hypometabolism as a key driver of HSC dysfunction with age and establish autophagy as a targetable node to reset oHSC regenerative capacity.
    Keywords:  aging; autophagy; hematopoietic stem cells; inflammation; metabolism; regeneration
    DOI:  https://doi.org/10.1016/j.stem.2024.04.020
  4. Cell Rep Med. 2024 May 06. pii: S2666-3791(24)00250-7. [Epub ahead of print] 101558
    Age-specific induction of mutant p53 drives clonal hematopoiesis and acute myeloid leukemia in adult mice.
    Rasoul Pourebrahim, Rafael Heinz Montoya, Hiroki Akiyama, Lauren Ostermann, Shayuan Khazaei, Muharrem Muftuoglu, Natalia Baran, Ran Zhao, Tom Lesluyes, Bin Liu, Joseph D Khoury, Mihai Gagea, Peter Van Loo, Michael Andreeff.
      The investigation of the mechanisms behind p53 mutations in acute myeloid leukemia (AML) has been limited by the lack of suitable mouse models, which historically have resulted in lymphoma rather than leukemia. This study introduces two new AML mouse models. One model induces mutant p53 and Mdm2 haploinsufficiency in early development, showing the role of Mdm2 in myeloid-biased hematopoiesis and AML predisposition, independent of p53. The second model mimics clonal hematopoiesis by inducing mutant p53 in adult hematopoietic stem cells, demonstrating that the timing of p53 mutation determines AML vs. lymphoma development. In this context, age-related changes in hematopoietic stem cells (HSCs) collaborate with mutant p53 to predispose toward myeloid transformation rather than lymphoma development. Our study unveils new insights into the cooperative impact of HSC age, Trp53 mutations, and Mdm2 haploinsufficiency on clonal hematopoiesis and the development of myeloid malignancies.
    Keywords:  Mdm2 haploinsufficiency; TP53 mutations; acute myeloid leukemia; cholesterol biosynthesis; clonal hematopoiesis; hematopoietic stem cells; mevalonate pathway; mouse model; myeloid-biased hematopoiesis
    DOI:  https://doi.org/10.1016/j.xcrm.2024.101558
  5. Exp Hematol. 2024 May 11. pii: S0301-472X(24)00092-4. [Epub ahead of print] 104233
    Complexities of modelling the bone marrow microenvironment to facilitate haematopoietic research.
    Caroline Busch, Kudzai Nyamondo, Helen Wheadon.
      Haematopoiesis occurs in the bone marrow (BM), within a specialised microenvironment referred to as the stem cell niche, where the haematopoietic stem cells (HSCs) reside and are regulated for quiescence, self-renewal and differentiation through intrinsic and extrinsic mechanisms. The BM contains at least two distinctive HSC supportive niches: an endosteal osteoblastic niche, which supports quiescence and self-renewal and a more vascular/peri-sinusoidal niche that promotes proliferation and differentiation. Both associate with supporting mesenchymal stromal cells (MSCs). Within the more hypoxic osteoblastic niche, HSCs specifically interact with the osteoblasts that line the endosteal surface, which secrete several important HSC quiescence and maintenance regulatory factors. In vivo imaging indicates that the HSCs and progenitors located further away, in the vicinity of sinusoidal endothelial cells, are more proliferative. Here HSCs interact with endothelial cells via specific cell adhesion molecules. Endothelial cells also secrete several factors important for HSC homeostasis and proliferation. In addition, HSCs and MSCs are embedded within the extracellular matrix (ECM), an important network of proteins such as collagen, elastin, laminin, proteoglycans, vitronectin and fibronectin. The ECM provides mechanical characteristics such as stiffness and elasticity important for cell behaviour regulation. ECM proteins are also able to bind, sequester, display and distribute growth factors across the BM, thus directly affecting stem cell fate and regulation of haematopoiesis. These important physical and chemical features of the BM require careful consideration when creating three dimensional models of the BM.
    Keywords:  Bone marrow microenvironment; Extracellular matrix; Haematopoietic stem cell; Mesenchymal stem cell
    DOI:  https://doi.org/10.1016/j.exphem.2024.104233
  6. Int J Hematol. 2024 May 15.
    Metabolic dependencies of acute myeloid leukemia stem cells.
    Xiangguo Shi, Mengdie Feng, Daisuke Nakada.
      Acute myeloid leukemia (AML) is a heterogeneous hematologic malignancy primarily driven by an immature population of AML cells termed leukemia stem cells (LSCs) that are implicated in AML development, chemoresistance, and relapse. An emerging area of research in AML focuses on identifying and targeting the aberrant metabolism in LSCs. Dysregulated metabolism is involved in sustaining functional properties of LSCs, impeding myeloid differentiation, and evading programmed cell death, both in the process of leukemogenesis and in response to chemotherapy. This review discusses recent discoveries regarding the aberrant metabolic processes of AML LSCs that have begun to change the therapeutic landscape of AML.
    Keywords:  Acute myeloid leukemia; Leukemia stem cell; Metabolic regulation
    DOI:  https://doi.org/10.1007/s12185-024-03789-x
  7. Stem Cell Res Ther. 2024 May 15. 15(1): 142
    Progresses in overcoming the limitations of in vitro erythropoiesis using human induced pluripotent stem cells.
    Hyeonwoo Ju, Yeowon Sohn, Yoojun Nam, Yeri Alice Rim.
      Researchers have attempted to generate transfusable oxygen carriers to mitigate RBC supply shortages. In vitro generation of RBCs using stem cells such as hematopoietic stem and progenitor cells (HSPCs), embryonic stem cells (ESCs), and induced pluripotent stem cells (iPSCs) has shown promise. Specifically, the limited supplies of HSPCs and ethical issues with ESCs make iPSCs the most promising candidate for in vitro RBC generation. However, researchers have encountered some major challenges when using iPSCs to produce transfusable RBC products, such as enucleation and RBC maturation. In addition, it has proven difficult to manufacture these products on a large scale. In this review, we provide a brief overview of erythropoiesis and examine endeavors to recapitulate erythropoiesis in vitro using various cell sources. Furthermore, we explore the current obstacles and potential solutions aimed at enabling the large-scale production of transfusable RBCs in vitro.
    Keywords:  Bioreactors; Enucleation; Erythropoiesis; Induced pluripotent stem cell; Red blood cell
    DOI:  https://doi.org/10.1186/s13287-024-03754-9
  8. Elife. 2024 May 17. pii: RP95815. [Epub ahead of print]13
    PI3K/HSCB axis facilitates FOG1 nuclear translocation to promote erythropoiesis and megakaryopoiesis.
    Gang Liu, Yunxuan Hou, Xin Jin, Yixue Zhang, Chaoyue Sun, Chengquan Huang, Yujie Ren, Jianmin Gao, Xiuli Wang, Xiumei Jiang.
      Erythropoiesis and megakaryopoiesis are stringently regulated by signaling pathways. However, the precise molecular mechanisms through which signaling pathways regulate key transcription factors controlling erythropoiesis and megakaryopoiesis remain partially understood. Herein, we identified heat shock cognate B (HSCB), which is well known for its iron-sulfur cluster delivery function, as an indispensable protein for friend of GATA 1 (FOG1) nuclear translocation during erythropoiesis of K562 human erythroleukemia cells and cord-blood-derived human CD34+CD90+hematopoietic stem cells (HSCs), as well as during megakaryopoiesis of the CD34+CD90+HSCs. Mechanistically, HSCB could be phosphorylated by phosphoinositol-3-kinase (PI3K) to bind with and mediate the proteasomal degradation of transforming acidic coiled-coil containing protein 3 (TACC3), which otherwise detained FOG1 in the cytoplasm, thereby facilitating FOG1 nuclear translocation. Given that PI3K is activated during both erythropoiesis and megakaryopoiesis, and that FOG1 is a key transcription factor for these processes, our findings elucidate an important, previously unrecognized iron-sulfur cluster delivery independent function of HSCB in erythropoiesis and megakaryopoiesis.
    Keywords:  FOG1 nuclear translocation; HSCB; PI3K; cell biology; erythropoiesis; human; megakaryopoiesis
    DOI:  https://doi.org/10.7554/eLife.95815
  9. bioRxiv. 2024 May 05. pii: 2024.05.05.592590. [Epub ahead of print]
    PROTAC-induced Protein Functional Dynamics in Targeted Protein Degradation.
    Kingsley Y Wu, Ta I Hung, Chia-En A Chang.
      PROteolysis TArgeting Chimeras (PROTACs) are small molecules that induce target protein degradation via the ubiquitin-proteasome system. PROTACs recruit the target protein and E3 ligase; a critical first step is forming a ternary complex. However, while the formation a ternary complex is crucial, it may not always guarantee successful protein degradation. The dynamics of the PROTAC-induced degradation complex play a key role in ubiquitination and subsequent degradation. In this study, we computationally modelled protein complex structures and dynamics associated with a series of PROTACs featuring different linkers to investigate why these PROTACs, all of which formed ternary complexes with Cereblon (CRBN) E3 ligase and the target protein bromodomain-containing protein 4 (BRD4 BD1 ), exhibited varying degrees of degradation potency. We constructed the degradation machinery complexes with Culling-Ring Ligase 4A (CRL4A) E3 ligase scaffolds. Through atomistic molecular dynamics simulations, we illustrated how PROTAC-dependent protein dynamics facilitate the arrangement of surface lysine residues of BRD4 BD1 into the catalytic pocket of E2/ubiquitin for ubiquitination. Despite featuring identical warheads in this PROTAC series, the linkers were found to affect the residue-interaction networks, and thus governing the essential motions of the entire degradation machine for ubiquitination. These findings offer a dynamic perspective on ligand-induced protein degradation, providing insights to guide future PROTAC design endeavors.
    DOI:  https://doi.org/10.1101/2024.05.05.592590
  10. Nat Commun. 2024 May 16. 15(1): 4139
    Bone marrow stromal cells induce chromatin remodeling in multiple myeloma cells leading to transcriptional changes.
    Moritz Binder, Raphael E Szalat, Srikanth Talluri, Mariateresa Fulciniti, Hervé Avet-Loiseau, Giovanni Parmigiani, Mehmet K Samur, Nikhil C Munshi.
      The natural history of multiple myeloma is characterized by its localization to the bone marrow and its interaction with bone marrow stromal cells. The bone marrow stromal cells provide growth and survival signals, thereby promoting the development of drug resistance. Here, we show that the interaction between bone marrow stromal cells and myeloma cells (using human cell lines) induces chromatin remodeling of cis-regulatory elements and is associated with changes in the expression of genes involved in the cell migration and cytokine signaling. The expression of genes involved in these stromal interactions are observed in extramedullary disease in patients with myeloma and provides the rationale for survival of myeloma cells outside of the bone marrow microenvironment. Expression of these stromal interaction genes is also observed in a subset of patients with newly diagnosed myeloma and are akin to the transcriptional program of extramedullary disease. The presence of such adverse stromal interactions in newly diagnosed myeloma is associated with accelerated disease dissemination, predicts the early development of therapeutic resistance, and is of independent prognostic significance. These stromal cell induced transcriptomic and epigenomic changes both predict long-term outcomes and identify therapeutic targets in the tumor microenvironment for the development of novel therapeutic approaches.
    DOI:  https://doi.org/10.1038/s41467-024-47793-5
  11. Nat Commun. 2024 May 15. 15(1): 4114
    IRE1α determines ferroptosis sensitivity through regulation of glutathione synthesis.
    Dadi Jiang, Youming Guo, Tianyu Wang, Liang Wang, Yuelong Yan, Ling Xia, Rakesh Bam, Zhifen Yang, Hyemin Lee, Takao Iwawaki, Boyi Gan, Albert C Koong.
      Cellular sensitivity to ferroptosis is primarily regulated by mechanisms mediating lipid hydroperoxide detoxification. We show that inositol-requiring enzyme 1 (IRE1α), an endoplasmic reticulum (ER) resident protein critical for the unfolded protein response (UPR), also determines cellular sensitivity to ferroptosis. Cancer and normal cells depleted of IRE1α gain resistance to ferroptosis, while enhanced IRE1α expression promotes sensitivity to ferroptosis. Mechanistically, IRE1α's endoribonuclease activity cleaves and down-regulates the mRNA of key glutathione biosynthesis regulators glutamate-cysteine ligase catalytic subunit (GCLC) and solute carrier family 7 member 11 (SLC7A11). This activity of IRE1α is independent of its role in regulating the UPR and is evolutionarily conserved. Genetic deficiency and pharmacological inhibition of IRE1α have similar effects in inhibiting ferroptosis and reducing renal ischemia-reperfusion injury in mice. Our findings reveal a previously unidentified role of IRE1α to regulate ferroptosis and suggests inhibition of IRE1α as a promising therapeutic strategy to mitigate ferroptosis-associated pathological conditions.
    DOI:  https://doi.org/10.1038/s41467-024-48330-0
  12. Neoplasia. 2024 May 16. pii: S1476-5586(24)00045-9. [Epub ahead of print]53 101003
    Extracellular signals induce dynamic ER remodeling through αTAT1-dependent microtubule acetylation.
    Hannah R Ortiz, Paola Cruz Flores, Julia Podgorski, Aaron Ramonett, Tasmia Ahmed, Nadine Hempel, Pascale G Charest, Nathan A Ellis, Paul R Langlais, William R Montfort, Karthikeyan Mythreye, Sanjay Kumar, Nam Y Lee.
      Dynamic changes in the endoplasmic reticulum (ER) morphology are central to maintaining cellular homeostasis. Microtubules (MT) facilitate the continuous remodeling of the ER network into sheets and tubules by coordinating with many ER-shaping protein complexes, although how this process is controlled by extracellular signals remains unknown. Here we report that TAK1, a kinase responsive to various growth factors and cytokines including TGF-β and TNF-α, triggers ER tubulation by activating αTAT1, an MT-acetylating enzyme that enhances ER-sliding. We show that this TAK1/αTAT1-dependent ER remodeling promotes cell survival by actively downregulating BOK, an ER membrane-associated proapoptotic effector. While BOK is normally protected from degradation when complexed with IP3R, it is rapidly degraded upon their dissociation during the ER sheets-to-tubules conversion. These findings demonstrate a distinct mechanism of ligand-induced ER remodeling and suggest that the TAK1/αTAT1 pathway may be a key target in ER stress and dysfunction.
    Keywords:  Alpha TAT1; BOK; Endoplasmic reticulum; Microtubules; TAK1; TGF-beta
    DOI:  https://doi.org/10.1016/j.neo.2024.101003
  13. Nat Biotechnol. 2024 May 14.
    Generation of allogeneic CAR-NKT cells from hematopoietic stem and progenitor cells using a clinically guided culture method.
    Yan-Ruide Li, Yang Zhou, Jiaji Yu, Yu Jeong Kim, Miao Li, Derek Lee, Kuangyi Zhou, Yuning Chen, Yichen Zhu, Yu-Chen Wang, Zhe Li, Yanqi Yu, Zachary Spencer Dunn, Wenbin Guo, Xinjian Cen, Tiffany Husman, Aarushi Bajpai, Adam Kramer, Matthew Wilson, Ying Fang, Jie Huang, Shuo Li, Yonggang Zhou, Yuchong Zhang, Zoe Hahn, Enbo Zhu, Feiyang Ma, Calvin Pan, Aldons J Lusis, Jin J Zhou, Christopher S Seet, Donald B Kohn, Pin Wang, Xianghong Jasmine Zhou, Matteo Pellegrini, Benjamin R Puliafito, Sarah M Larson, Lili Yang.
      Cancer immunotherapy with autologous chimeric antigen receptor (CAR) T cells faces challenges in manufacturing and patient selection that could be avoided by using 'off-the-shelf' products, such as allogeneic CAR natural killer T (AlloCAR-NKT) cells. Previously, we reported a system for differentiating human hematopoietic stem and progenitor cells into AlloCAR-NKT cells, but the use of three-dimensional culture and xenogeneic feeders precluded its clinical application. Here we describe a clinically guided method to differentiate and expand IL-15-enhanced AlloCAR-NKT cells with high yield and purity. We generated AlloCAR-NKT cells targeting seven cancers and, in a multiple myeloma model, demonstrated their antitumor efficacy, expansion and persistence. The cells also selectively depleted immunosuppressive cells in the tumor microenviroment and antagonized tumor immune evasion via triple targeting of CAR, TCR and NK receptors. They exhibited a stable hypoimmunogenic phenotype associated with epigenetic and signaling regulation and did not induce detectable graft versus host disease or cytokine release syndrome. These properties of AlloCAR-NKT cells support their potential for clinical translation.
    DOI:  https://doi.org/10.1038/s41587-024-02226-y
  14. Blood Adv. 2024 May 13. pii: bloodadvances.2022008144. [Epub ahead of print]
    Deep PIM kinase substrate profiling reveals new rational co-therapeutic strategies for acute myeloid leukemia.
    Tejashree Joglekar, Alexander Chin, Alin Voskanian-Kordi, Baek Seungchul, Azim Raja, Apurv Rege, Weiliang Huang, Maureen A Kane, Marikki Laiho, Thomas Webb, Xiaoxuan Fan, Michael Rubenstein, Charles J Bieberich, Xiang Li.
      Provirus integration site for Moloney murine leukemia virus (PIM) family serine/threonine kinases perform pro-tumorigenic functions in hematologic malignancies and solid tumors by phosphorylating substrates involved in tumor metabolism, cell survival, metastasis, inflammation, and immune cell invasion. However, a comprehensive understanding of PIM kinase functions is currently lacking. Multiple small molecule PIM kinase inhibitors are currently being evaluated as co-therapeutics in cancer patients. To further illuminate PIM kinase functions in cancer, we deeply profiled PIM1 substrates using the reverse in-gel kinase assay to identify downstream cellular processes targetable with small molecules. Pathway analyses of putative PIM substrates nominated RNA splicing and rRNA processing as PIM-regulated cellular processes. PIM inhibition elicited reproducible splicing changes in PIM-inhibitor-responsive acute myeloid leukemia (AML) cell lines. PIM inhibitors synergized with splicing modulators targeting splicing factor 3b subunit 1 (SF3B1) and serine-arginine protein kinase 1 (SRPK1) to kill AML cells. PIM inhibition also altered rRNA processing, and PIM inhibitors synergized with an RNA polymerase I inhibitor to kill AML cells and block AML tumor growth. These data demonstrate that deep kinase substrate knowledge can illuminate unappreciated kinase functions, nominating synergistic co-therapeutic strategies. This approach may expand the co-therapeutic armamentarium to overcome kinase-inhibitor resistant disease that limits durable responses in malignant disease.
    DOI:  https://doi.org/10.1182/bloodadvances.2022008144
  15. Nat Commun. 2024 May 17. 15(1): 3812
    High-throughput mechanical phenotyping and transcriptomics of single cells.
    Akifumi Shiomi, Taikopaul Kaneko, Kaori Nishikawa, Arata Tsuchida, Takashi Isoshima, Mayuko Sato, Kiminori Toyooka, Kentaro Doi, Hidekazu Nishikii, Hirofumi Shintaku.
      The molecular system regulating cellular mechanical properties remains unexplored at single-cell resolution mainly due to a limited ability to combine mechanophenotyping with unbiased transcriptional screening. Here, we describe an electroporation-based lipid-bilayer assay for cell surface tension and transcriptomics (ELASTomics), a method in which oligonucleotide-labelled macromolecules are imported into cells via nanopore electroporation to assess the mechanical state of the cell surface and are enumerated by sequencing. ELASTomics can be readily integrated with existing single-cell sequencing approaches and enables the joint study of cell surface mechanics and underlying transcriptional regulation at an unprecedented resolution. We validate ELASTomics via analysis of cancer cell lines from various malignancies and show that the method can accurately identify cell types and assess cell surface tension. ELASTomics enables exploration of the relationships between cell surface tension, surface proteins, and transcripts along cell lineages differentiating from the haematopoietic progenitor cells of mice. We study the surface mechanics of cellular senescence and demonstrate that RRAD regulates cell surface tension in senescent TIG-1 cells. ELASTomics provides a unique opportunity to profile the mechanical and molecular phenotypes of single cells and can dissect the interplay among these in a range of biological contexts.
    DOI:  https://doi.org/10.1038/s41467-024-48088-5
  16. Elife. 2024 May 16. pii: RP88275. [Epub ahead of print]12
    Long-term hematopoietic transfer of the anti-cancer and lifespan-extending capabilities of a genetically engineered blood system by transplantation of bone marrow mononuclear cells.
    Jing-Ping Wang, Chun-Hao Hung, Yae-Huei Liou, Ching-Chen Liu, Kun-Hai Yeh, Keh-Yang Wang, Zheng-Sheng Lai, Biswanath Chatterjee, Tzu-Chi Hsu, Tung-Liang Lee, Yu-Chiau Shyu, Pei-Wen Hsiao, Liuh-Yow Chen, Trees-Juen Chuang, Chen-Hsin Albert Yu, Nan-Shih Liao, C-K James Shen.
      A causal relationship exists among the aging process, organ decay and disfunction, and the occurrence of various diseases including cancer. A genetically engineered mouse model, termed Klf1K74R/K74R or Klf1(K74R), carrying mutation on the well-conserved sumoylation site of the hematopoietic transcription factor KLF1/EKLF has been generated that possesses extended lifespan and healthy characteristics, including cancer resistance. We show that the healthy longevity characteristics of the Klf1(K74R) mice, as exemplified by their higher anti-cancer capability, are likely gender-, age-, and genetic background-independent. Significantly, the anti-cancer capability, in particular that against melanoma as well as hepatocellular carcinoma, and lifespan-extending property of Klf1(K74R) mice, could be transferred to wild-type mice via transplantation of their bone marrow mononuclear cells at a young age of the latter. Furthermore, NK(K74R) cells carry higher in vitro cancer cell-killing ability than wild-type NK cells. Targeted/global gene expression profiling analysis has identified changes in the expression of specific proteins, including the immune checkpoint factors PDCD and CD274, and cellular pathways in the leukocytes of the Klf1(K74R) that are in the directions of anti-cancer and/or anti-aging. This study demonstrates the feasibility of developing a transferable hematopoietic/blood system for long-term anti-cancer and, potentially, for anti-aging.
    Keywords:  EKLF/ KLF1; aging; anti-cancer; bone marrow transplantation; leukocytes; mouse; regenerative medicine; stem cells
    DOI:  https://doi.org/10.7554/eLife.88275
  17. Cell Rep Med. 2024 May 08. pii: S2666-3791(24)00264-7. [Epub ahead of print] 101572
    CD37 is a safe chimeric antigen receptor target to treat acute myeloid leukemia.
    Benjamin Caulier, Sandy Joaquina, Pascal Gelebart, Tara Helén Dowling, Fatemeh Kaveh, Moritz Thomas, Luka Tandaric, Patrik Wernhoff, Niveditha Umesh Katyayini, Cara Wogsland, May Eriksen Gjerstad, Yngvar Fløisand, Gunnar Kvalheim, Carsten Marr, Sebastian Kobold, Jorrit M Enserink, Bjørn Tore Gjertsen, Emmet McCormack, Else Marit Inderberg, Sébastien Wälchli.
      Acute myeloid leukemia (AML) is characterized by the accumulation of immature myeloid cells in the bone marrow and the peripheral blood. Nearly half of the AML patients relapse after standard induction therapy, and new forms of therapy are urgently needed. Chimeric antigen receptor (CAR) T therapy has so far not been successful in AML due to lack of efficacy and safety. Indeed, the most attractive antigen targets are stem cell markers such as CD33 or CD123. We demonstrate that CD37, a mature B cell marker, is expressed in AML samples, and its presence correlates with the European LeukemiaNet (ELN) 2017 risk stratification. We repurpose the anti-lymphoma CD37CAR for the treatment of AML and show that CD37CAR T cells specifically kill AML cells, secrete proinflammatory cytokines, and control cancer progression in vivo. Importantly, CD37CAR T cells display no toxicity toward hematopoietic stem cells. Thus, CD37 is a promising and safe CAR T cell AML target.
    Keywords:  AML; CAR T cell; CD37; acute myeloid leukemia; chimeric antigen receptor; hematopoietic stem cell; immunotherapy; patient-derived xenograft
    DOI:  https://doi.org/10.1016/j.xcrm.2024.101572
  18. Methods Mol Biol. 2024 ;2804 141-162
    A Guide to the Quantitation of Protein Secretion Dynamics at the Single-Cell Level.
    Nathan Aymerich, Olivia T M Bucheli, Kevin Portmann, Klaus Eyer, Jean Baudry.
      Protein secretion is a key cellular functionality, particularly in immunology, where cells can display large heterogeneity in this crucial activity in addition to binary secretion behavior. However, few methods enable quantitative secretion rate measurements at the single-cell level, and these methods are mostly based on microfluidics systems. Here, we describe such a microfluidic single-cell method for precisely measuring protein secretion rates in detail, building on the published droplet-based microfluidic platform DropMap. We give an updated, detailed guide toward quantifying protein secretion rates, discussing its setup and limitations. We illustrate the protocol on two key immunological analytes, immunoglobulin G, and interferon-γ.
    Keywords:  Droplet microfluidics; Immune cells; Protein secretion; Quantification; Secretion rates; Single-cell analysis
    DOI:  https://doi.org/10.1007/978-1-0716-3850-7_9
  19. Leukemia. 2024 May 11.
    TIF1β activates leukemic transcriptional program in HSCs and promotes BCR::ABL1-induced myeloid leukemia.
    Mariko Morii, Sho Kubota, Mihoko Iimori, Takako Yokomizo-Nakano, Ai Hamashima, Jie Bai, Akiho Nishimura, Masayoshi Tasaki, Yukio Ando, Kimi Araki, Goro Sashida.
      TIF1β/KAP1/TRIM28, a chromatin modulator, both represses and activates the transcription of genes in normal and malignant cells. Analyses of datasets on leukemia patients revealed that the expression level of TIF1β was increased in patients with chronic myeloid leukemia at the blast crisis and acute myeloid leukemia. We generated a BCR::ABL1 conditional knock-in (KI) mouse model, which developed aggressive myeloid leukemia, and demonstrated that the deletion of the Tif1β gene inhibited the progression of myeloid leukemia and showed longer survival than that in BCR::ABL1 KI mice, suggesting that Tif1β drove the progression of BCR::ABL1-induced leukemia. In addition, the deletion of Tif1β sensitized BCR::ABL1 KI leukemic cells to dasatinib. The deletion of Tif1β decreased the expression levels of TIF1β-target genes and chromatin accessibility peaks enriched with the Fosl1-binding motif in BCR::ABL1 KI stem cells. TIF1β directly bound to the promoters of proliferation genes, such as FOSL1, in human BCR::ABL1 cells, in which TIF1β and FOSL1 bound to adjacent regions of chromatin. Since the expression of Fosl1 was critical for the enhanced growth of BCR::ABL1 KI cells, Tif1β and Fosl1 interacted to activate the leukemic transcriptional program in and cellular function of BCR::ABL1 KI stem cells and drove the progression of myeloid leukemia.
    DOI:  https://doi.org/10.1038/s41375-024-02276-w
  20. Cell. 2024 May 08. pii: S0092-8674(24)00413-6. [Epub ahead of print]
    An age-progressive platelet differentiation path from hematopoietic stem cells causes exacerbated thrombosis.
    Donna M Poscablo, Atesh K Worthington, Stephanie Smith-Berdan, Marcel G E Rommel, Bryce A Manso, Reheman Adili, Lydia Mok, Roman E Reggiardo, Taylor Cool, Raana Mogharrab, Jenna Myers, Steven Dahmen, Paloma Medina, Anna E Beaudin, Scott W Boyer, Michael Holinstat, Vanessa D Jonsson, E Camilla Forsberg.
      Platelet dysregulation is drastically increased with advanced age and contributes to making cardiovascular disorders the leading cause of death of elderly humans. Here, we reveal a direct differentiation pathway from hematopoietic stem cells into platelets that is progressively propagated upon aging. Remarkably, the aging-enriched platelet path is decoupled from all other hematopoietic lineages, including erythropoiesis, and operates as an additional layer in parallel with canonical platelet production. This results in two molecularly and functionally distinct populations of megakaryocyte progenitors. The age-induced megakaryocyte progenitors have a profoundly enhanced capacity to engraft, expand, restore, and reconstitute platelets in situ and upon transplantation and produce an additional platelet population in old mice. The two pools of co-existing platelets cause age-related thrombocytosis and dramatically increased thrombosis in vivo. Strikingly, aging-enriched platelets are functionally hyper-reactive compared with the canonical platelet populations. These findings reveal stem cell-based aging as a mechanism for platelet dysregulation and age-induced thrombosis.
    Keywords:  aging; cardiovascular disorder; genetic lineage tracing; hematopoietic stem cells; megakaryocyte progenitor cells; platelet differentiation pathway; platelet dysregulation; thrombocytosis; thrombosis; transplantation
    DOI:  https://doi.org/10.1016/j.cell.2024.04.018
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