bims-scepro 2024-07-14 papers

bims-scepro

Biomed News

on Stem cell proteostasis

Issue of 2024–07–14
thirty-one papers selected by
William Grey, University of York

Hematopoietic Stem Cells and Their Niche in Bone Marrow.
Bioengineered niches that recreate physiological extracellular matrix organisation to support long-term haematopoietic stem cells.
Endomucin marks quiescent long-term multi-lineage repopulating hematopoietic stem cells and is essential for their transendothelial migration.
tRNA m1A modification regulate HSC maintenance and self-renewal via mTORC1 signaling.
In Fanconi Anemia, impaired accumulation of bone marrow neutrophils during emergency granulopoiesis induces hematopoietic stem cell stress.
Silencing endomucin in bone marrow sinusoids improves hematopoietic stem and progenitor cell homing during transplantation.
Automated single-cell proteomics providing sufficient proteome depth to study complex biology beyond cell type classifications.
How aging influences the gut-bone marrow axis and alters hematopoietic stem cell regulation.
Inhibitors of the tyrosine kinases FMS-like tyrosine kinase-3 and WEE1 induce apoptosis and DNA damage synergistically in acute myeloid leukemia cells.
New frameworks for hematopoiesis derived from single-cell genomics.
Anti-TIM3 chimeric antigen receptor-natural killer cells preferentially target primitive acute myeloid leukemia cells with minimal fratricide and exhaustion.
Emerging role of glutathione peroxidase 4 in myeloid cell lineage development and acute myeloid leukemia.
CDK6: HSC fate and cell cycle nexus?
International Society for Cell & Gene Therapy Stem Cell Engineering Committee report on the current state of hematopoietic stem and progenitor cell-based genomic therapies and the challenges faced.
Differential inflammatory conditioning of the bone marrow by acute myeloid leukemia and its impact on progression.
Hematopoietic stem and progenitor cell membrane-coated vesicles for bone marrow-targeted leukaemia drug delivery.
Are We There Yet? Assessing the Readiness of Single-Cell Proteomics to Answer Biological Hypotheses.
RNF149 Destabilizes IFNGR1 in Macrophages to Favor Post-Infarction Cardiac Repair.
Targeting of human cancer stem cells predicts efficacy and toxicity of FDA-approved oncology drugs.
Aging-related changes in the mechanical properties of single cells.
Dual-localized PPTC7 limits mitophagy through proximal and dynamic interactions with BNIP3 and NIX.
Functional characterization of native Piezo1 as calcium and magnesium influx pathway in human myeloid leukemia cells.
Heterogeneous stiffness of the bone marrow microenvironment regulates the fate decision of haematopoietic stem and progenitor cells.
Erythroid-intrinsic activation of TLR8 impairs erythropoiesis in inherited anemia.
Plasmacytoid dendritic cells control homeostasis of megakaryopoiesis.
Targeting cancer stem cells in multiple myeloma.
A 3D micro-printed single cell micro-niche with asymmetric niche signals - An in vitro model for asymmetric cell division study.
Epigenetic alterations affecting hematopoietic regulatory networks as drivers of mixed myeloid/lymphoid leukemia.
FAM110A promotes mitotic spindle formation by linking microtubules with actin cytoskeleton.
Distinct phases of cellular signaling revealed by time-resolved protein synthesis.
Cell-intrinsic mechanical regulation of plasma membrane accumulation at the cytokinetic furrow.

Int J Mol Sci. 2024 Jun 21. pii: 6837. [Epub ahead of print]25(13):

Hematopoietic Stem Cells and Their Niche in Bone Marrow.

Munju Kwon, Byoung Soo Kim, Sik Yoon, Sae-Ock Oh, Dongjun Lee.

  Extensive research has explored the functional correlation between stem cells and progenitor cells, particularly in blood. Hematopoietic stem cells (HSCs) can self-renew and regenerate tissues within the bone marrow, while stromal cells regulate tissue function. Recent studies have validated the role of mammalian stem cells within specific environments, providing initial empirical proof of this functional phenomenon. The interaction between bone and blood has always been vital to the function of the human body. It was initially proposed that during evolution, mammalian stem cells formed a complex relationship with the surrounding microenvironment, known as the niche. Researchers are currently debating the significance of molecular-level data to identify individual stromal cell types due to incomplete stromal cell mapping. Obtaining these data can help determine the specific activities of HSCs in bone marrow. This review summarizes key topics from previous studies on HSCs and their environment, discussing current and developing concepts related to HSCs and their niche in the bone marrow.

Keywords:  bone marrow microenvironment; hematopoietic progenitor cells; hematopoietic stem cells; niche

DOI:  https://doi.org/10.3390/ijms25136837
Nat Commun. 2024 Jul 10. 15(1): 5791

Bioengineered niches that recreate physiological extracellular matrix organisation to support long-term haematopoietic stem cells.

Hannah Donnelly, Ewan Ross, Yinbo Xiao, Rio Hermantara, Aqeel F Taqi, W Sebastian Doherty-Boyd, Jennifer Cassels, Penelope M Tsimbouri, Karen M Dunn, Jodie Hay, Annie Cheng, R M Dominic Meek, Nikhil Jain, Christopher West, Helen Wheadon, Alison M Michie, Bruno Peault, Adam G West, Manuel Salmeron-Sanchez, Matthew J Dalby.

Long-term reconstituting haematopoietic stem cells (LT-HSCs) are used to treat blood disorders via stem cell transplantation. The very low abundance of LT-HSCs and their rapid differentiation during in vitro culture hinders their clinical utility. Previous developments using stromal feeder layers, defined media cocktails, and bioengineering have enabled HSC expansion in culture, but of mostly short-term HSCs and progenitor populations at the expense of naive LT-HSCs. Here, we report the creation of a bioengineered LT-HSC maintenance niche that recreates physiological extracellular matrix organisation, using soft collagen type-I hydrogels to drive nestin expression in perivascular stromal cells (PerSCs). We demonstrate that nestin, which is expressed by HSC-supportive bone marrow stromal cells, is cytoprotective and, via regulation of metabolism, is important for HIF-1α expression in PerSCs. When CD34+ve HSCs were added to the bioengineered niches comprising nestin/HIF-1α expressing PerSCs, LT-HSC numbers were maintained with normal clonal and in vivo reconstitution potential, without media supplementation. We provide proof-of-concept that our bioengineered niches can support the survival of CRISPR edited HSCs. Successful editing of LT-HSCs ex vivo can have potential impact on the treatment of blood disorders.

DOI: https://doi.org/10.1038/s41467-024-50054-0
Cell Rep. 2024 Jul 10. pii: S2211-1247(24)00804-0. [Epub ahead of print]43(7): 114475

Endomucin marks quiescent long-term multi-lineage repopulating hematopoietic stem cells and is essential for their transendothelial migration.

Sophia Engelhard, Montserrat Estruch, Shuyu Qin, Christoph A Engelhard, Francisco G Rodriguez-Gonzalez, Martine Drilsvik, Javier Martin-Gonzalez, Jeng-Wei Lu, David Bryder, Claus Nerlov, Joachim Weischenfeldt, Kristian Reckzeh, Kim Theilgaard-Mönch.

  Endomucin (EMCN) currently represents the only hematopoietic stem cell (HSC) marker expressed by both murine and human HSCs. Here, we report that EMCN+ long-term repopulating HSCs (LT-HSCs; CD150+CD48-LSK) have a higher long-term multi-lineage repopulating capacity compared to EMCN- LT-HSCs. Cell cycle analyses and transcriptional profiling demonstrated that EMCN+ LT-HSCs were more quiescent compared to EMCN- LT-HSCs. Emcn-/- and Emcn+/+ mice displayed comparable steady-state hematopoiesis, as well as frequencies, transcriptional programs, and long-term multi-lineage repopulating capacity of their LT-HSCs. Complementary functional analyses further revealed increased cell cycle entry upon treatment with 5-fluorouracil and reduced granulocyte colony-stimulating factor (GCSF) mobilization of Emcn-/- LT-HSCs, demonstrating that EMCN expression by LT-HSCs associates with quiescence in response to hematopoietic stress and is indispensable for effective LT-HSC mobilization. Transplantation of wild-type bone marrow cells into Emcn-/- or Emcn+/+ recipients demonstrated that EMCN is essential for endothelial cell-dependent maintenance/self-renewal of the LT-HSC pool and sustained blood cell production post-transplant.

Keywords:  CP: Stem cell research; bone marrow niche; cross-species HSC marker; endomucin; endothelial cells; hematopoietic stem cell; mobilization; transplantation

DOI:  https://doi.org/10.1016/j.celrep.2024.114475
Nat Commun. 2024 Jul 08. 15(1): 5706

tRNA m1A modification regulate HSC maintenance and self-renewal via mTORC1 signaling.

Hongna Zuo, Aiwei Wu, Mingwei Wang, Liquan Hong, Hu Wang.

Haematopoietic stem cells (HSCs) possess unique physiological adaptations to sustain blood cell production and cope with stress responses throughout life. To maintain these adaptations, HSCs rely on maintaining a tightly controlled protein translation rate. However, the mechanism of how HSCs regulate protein translation remains to be fully elucidated. In this study, we investigate the role of transfer RNA (tRNA) m1A58 'writer' proteins TRMT6 and TRMT61A in regulating HSCs function. Trmt6 deletion promoted HSC proliferation through aberrant activation of mTORC1 signaling. TRMT6-deficient HSCs exhibited an impaired self-renewal ability in competitive transplantation assay. Mechanistically, single cell RNA-seq analysis reveals that the mTORC1 signaling pathway is highly upregulated in HSC-enriched cell populations after Trmt6 deletion. m1A-tRNA-seq and Western blot analysis suggest that TRMT6 promotes methylation modification of specific tRNA and expression of TSC1, fine-tuning mTORC1 signaling levels. Furthermore, Pharmacological inhibition of the mTORC1 pathway rescued functional defect in TRMT6-deficient HSCs. To our knowledge, this study is the first to elucidate a mechanism by which TRMT6-TRMT61A complex-mediated tRNA-m1A58 modification regulates HSC homeostasis.

DOI: https://doi.org/10.1038/s41467-024-50110-9
J Biol Chem. 2024 Jul 09. pii: S0021-9258(24)02049-0. [Epub ahead of print] 107548

In Fanconi Anemia, impaired accumulation of bone marrow neutrophils during emergency granulopoiesis induces hematopoietic stem cell stress.

Liping Hu, Weiqi Huang, Bin Liu, Elizabeth A Eklund.

  Fanconi Anemia (FA) is an inherited disorder of DNA-repair due to mutation in one of 20+ interrelated genes that repair intra-strand DNA crosslinks and rescue collapsed or stalled replication forks. The most common hematologic abnormality in FA is anemia, but progression to bone marrow failure (BMF), clonal hematopoiesis, or acute myeloid leukemia (AML) may also occur. In prior studies, we found that Fanconi DNA-repair is required for successful emergency granulopoiesis; the process for rapid neutrophil production during the innate immune response. Specifically, Fancc-/- mice did not develop neutrophilia in response to emergency granulopoiesis stimuli, but instead exhibited apoptosis of bone marrow hematopoietic stem cells (HSCs) and differentiating neutrophils. Repeated emergency granulopoiesis challenges induced BMF in most Fancc-/- mice, with AML in survivors. In contrast, we found equivalent neutrophilia during emergency granulopoiesis in Fancc-/-Tp53+/- mice and wild type (WT) mice, without BMF. Since termination of emergency granulopoiesis is triggered by accumulation of bone marrow neutrophils, we hypothesize neutrophilia protects Fancc-/-Tp53+/- bone marrow from the stress of a sustained inflammation that is experienced by Fancc-/- mice. In the current work, we found that blocking neutrophil accumulation during emergency granulopoiesis led to BMF in Fancc-/-Tp53+/- mice, consistent with this hypothesis. Blocking neutrophilia during emergency granulopoiesis in Fancc-/-Tp53+/- mice (but not WT) impaired cell cycle checkpoint activity, also found in Fancc-/- mice. Mechanisms for loss of cell cycle checkpoints during infections challenges may define molecular markers of FA progression, or suggest therapeutic targets for bone marrow protection in this disorder.

Keywords:  DNA damage response; DNA repair; cell cycle; hematopoiesis; inflammasome; innate immunity; neutrophil

DOI:  https://doi.org/10.1016/j.jbc.2024.107548
Stem Cells. 2024 Jul 12. pii: sxae046. [Epub ahead of print]

Silencing endomucin in bone marrow sinusoids improves hematopoietic stem and progenitor cell homing during transplantation.

Yue Li, Miao Ren, Hu Li, Zuo Zhang, Ke Yuan, Yujin Huang, Shengnan Yuan, Wen Ju, Yuan He, Kailin Xu, Lingyu Zeng.

  Efficient homing of infused hematopoietic stem and progenitor cells (HSPCs) into the bone marrow (BM) is the prerequisite for successful hematopoietic stem cell transplantation. However, only a small part of infused HSPCs find their way to the BM niche. A better understanding of the mechanisms that facilitate HSPC homing will help to develop strategies to improve the initial HSPC engraftment and subsequent hematopoietic regeneration. Here, we show that irradiation upregulates the endomucin expression of endothelial cells in vivo and in vitro. Furthermore, depletion of endomucin in irradiated endothelial cells with short interfering RNA (siRNA) increases the HSPC-endothelial cell adhesion in vitro. To abrogate the endomucin of BM sinusoidal endothelial cells (BM-SECs) in vivo, we develop a siRNA-loaded bovine serum albumin nanoparticle for targeted delivery. Nanoparticle-mediated siRNA delivery successfully silences endomucin expression in BM-SECs and improves HSPC homing during transplantation. These results reveal that endomucin plays a critical role in HSPC homing during transplantation and that gene-based manipulation of BM-SEC endomucin in vivo can be exploited to improve the efficacy of HSPC transplantation.

Keywords:  HSPC; Hematopoietic stem cell transplantation; bone marrow sinusoidal endothelial cell; endomucin; homing; therapeutic siRNA

DOI:  https://doi.org/10.1093/stmcls/sxae046
Nat Commun. 2024 Jul 08. 15(1): 5707

Automated single-cell proteomics providing sufficient proteome depth to study complex biology beyond cell type classifications.

Claudia Ctortecka, Natalie M Clark, Brian W Boyle, Anjali Seth, D R Mani, Namrata D Udeshi, Steven A Carr.

The recent technological and computational advances in mass spectrometry-based single-cell proteomics have pushed the boundaries of sensitivity and throughput. However, reproducible quantification of thousands of proteins within a single cell remains challenging. To address some of those limitations, we present a dedicated sample preparation chip, the proteoCHIP EVO 96 that directly interfaces with the Evosep One. This, in combination with the Bruker timsTOF demonstrates double the identifications without manual sample handling and the newest generation timsTOF Ultra identifies up to 4000 with an average of 3500 protein groups per single HEK-293T without a carrier or match-between runs. Our workflow spans 4 orders of magnitude, identifies over 50 E3 ubiquitin-protein ligases, and profiles key regulatory proteins upon small molecule stimulation. This study demonstrates that the proteoCHIP EVO 96-based sample preparation with the timsTOF Ultra provides sufficient proteome depth to study complex biology beyond cell-type classifications.

DOI: https://doi.org/10.1038/s41467-024-49651-w
Heliyon. 2024 Jun 30. 10(12): e32831

How aging influences the gut-bone marrow axis and alters hematopoietic stem cell regulation.

Christopher Wells, Tristan Robertson, Prameet Sheth, Sheela Abraham.

  The gut microbiome has come to prominence across research disciplines, due to its influence on major biological systems within humans. Recently, a relationship between the gut microbiome and hematopoietic system has been identified and coined the gut-bone marrow axis. It is well established that the hematopoietic system and gut microbiome separately alter with age; however, the relationship between these changes and how these systems influence each other demands investigation. Since the hematopoietic system produces immune cells that help govern commensal bacteria, it is important to identify how the microbiome interacts with hematopoietic stem cells (HSCs). The gut microbiota has been shown to influence the development and outcomes of hematologic disorders, suggesting dysbiosis may influence the maintenance of HSCs with age. Short chain fatty acids (SCFAs), lactate, iron availability, tryptophan metabolites, bacterial extracellular vesicles, microbe associated molecular patterns (MAMPs), and toll-like receptor (TLR) signalling have been proposed as key mediators of communication across the gut-bone marrow axis and will be reviewed in this article within the context of aging.

Keywords:  Aging; Gut dysbiosis; Hematologic disorders; Hematopoietic stem cells; Inflammation; Microbiome; Stem cell regulation

DOI:  https://doi.org/10.1016/j.heliyon.2024.e32831
Biomed Pharmacother. 2024 Jul 05. pii: S0753-3322(24)00960-0. [Epub ahead of print]177 117076

Inhibitors of the tyrosine kinases FMS-like tyrosine kinase-3 and WEE1 induce apoptosis and DNA damage synergistically in acute myeloid leukemia cells.

Christoph Hieber, Al-Hassan M Mustafa, Sarah Neuroth, Sven Henninger, Hans-Peter Wollscheid, Joanna Zabkiewicz, Michelle Lazenby, Caroline Alvares, Siavosh Mahboobi, Falk Butter, Walburgis Brenner, Matthias Bros, Oliver H Krämer.

  Hyperactive FMS-like receptor tyrosine kinase-3 mutants with internal tandem duplications (FLT3-ITD) are frequent driver mutations of aggressive acute myeloid leukemia (AML). Inhibitors of FLT3 produce promising results in rationally designed cotreatment schemes. Since FLT3-ITD modulates DNA replication and DNA repair, valid anti-leukemia strategies could rely on a combined inhibition of FLT3-ITD and regulators of cell cycle progression and DNA integrity. These include the WEE1 kinase which controls cell cycle progression, nucleotide synthesis, and DNA replication origin firing. We investigated how pharmacological inhibition of FLT3 and WEE1 affected the survival and genomic integrity of AML cell lines and primary AML cells. We reveal that promising clinical grade and preclinical inhibitors of FLT3 and WEE1 synergistically trigger apoptosis in leukemic cells that express FLT3-ITD. An accumulation of single and double strand DNA damage precedes this process. Mass spectrometry-based proteomic analyses show that FLT3-ITD and WEE1 sustain the expression of the ribonucleotide reductase subunit RRM2, which provides dNTPs for DNA replication. Unlike their strong pro-apoptotic effects on leukemia cells with FLT3-ITD, inhibitors of FLT3 and WEE1 do not damage healthy human blood cells and murine hematopoietic stem cells. Thus, pharmacological inhibition of FLT3-ITD and WEE1 might become an improved, rationally designed therapeutic option.

Keywords:  AML; DNA replication stress; FLT3-ITD; Inhibitors; Synergism; WEE1

DOI:  https://doi.org/10.1016/j.biopha.2024.117076
Blood. 2024 Jul 10. pii: blood.2024024006. [Epub ahead of print]

New frameworks for hematopoiesis derived from single-cell genomics.

Ksenia Safina, Peter van Galen.

Recent advancements in single-cell genomics have enriched our understanding of hematopoiesis, providing intricate details about hematopoietic stem cell (HSC) biology, differentiation, and lineage commitment. Technological advancements have highlighted extensive heterogeneity of cell populations and continuity of differentiation routes. Nevertheless, intermediate 'attractor' states signify structure in stem and progenitor populations that link state transition dynamics to fate potential. We discuss how innovative model systems quantify lineage bias and how stress accelerates differentiation, thereby reducing fate plasticity compared to native hematopoiesis. We conclude by offering our perspective on the current model of hematopoiesis and discuss how a more precise understanding can translate to strategies that extend healthy hematopoiesis and prevent disease.

DOI: https://doi.org/10.1182/blood.2024024006
Exp Hematol Oncol. 2024 Jul 11. 13(1): 67

Anti-TIM3 chimeric antigen receptor-natural killer cells preferentially target primitive acute myeloid leukemia cells with minimal fratricide and exhaustion.

Phatchanat Klaihmon, Parinya Samart, Yon Rojanasakul, Surapol Issaragrisil, Sudjit Luanpitpong.

  Acute myeloid leukemia (AML) is an aggressive and genetically heterogeneous disease with poor clinical outcomes. Refractory AML is common, and relapse remains a major challenge, attributable to the presence of therapy-resistant leukemic stem cells (LSCs), which possess self-renewal and repopulating capability. Targeting LSCs is currently the most promising avenue for long-term management of AML. Likewise, chimeric antigen receptor (CAR)-natural killer (NK) cells have emerged as a promising alternative to CAR-T cells due to their intrinsic potential as off-the-shelf products and safer clinical profiles. Here, we introduced a third-generation CAR harboring TIM3 scFv, CD28, 4-1BB, and CD3ζ (CAR-TIM3) into human NK-92 cells, the only FDA-approved NK cell line for clinical trials. TIM3 was chosen as a target antigen owing to its differential expression in LSCs and normal hematopoietic stem/progenitor cells (HSPCs). The established CAR-TIM3 NK-92 cells effectively targeted TIM3 and displayed potent anti-tumor activity against various primitive AML cells, subsequently causing a reduction in leukemic clonogenic growth in vitro, while having minimal effects on HSPCs. CAR-TIM3 NK-92 cells significantly reduced leukemic burden in vivo and interestingly suppressed the engraftment of AML cells into the mouse liver and bone marrow. Surprisingly, we found that CAR-TIM3 NK-92 cells expressed relatively low surface TIM3, leading to a low fratricidal effect. As TIM3 and PD-1 are immune checkpoints involved in NK cell dysfunction, we further tested and found that CAR-TIM3 NK-92 cells are beneficial for alleviating NK cell exhaustion. Our findings highlight the potential application of CAR-TIM3 NK cells for cellular immunotherapy for TIM3+ AML.

Keywords:  Acute myeloid leukemia; Chimeric antigen receptor; Exhaustion; Immunotherapy; Natural killer cell; TIM3

DOI:  https://doi.org/10.1186/s40164-024-00534-2
Cell Mol Biol Lett. 2024 Jul 08. 29(1): 98

Emerging role of glutathione peroxidase 4 in myeloid cell lineage development and acute myeloid leukemia.

Patrick Auberger, Cécile Favreau, Coline Savy, Arnaud Jacquel, Guillaume Robert.

  Phospholipid Hydroperoxide Gluthatione Peroxidase also called Glutathione Peroxidase 4 is one of the 25 described human selenoproteins. It plays an essential role in eliminating toxic lipid hydroxy peroxides, thus inhibiting ferroptosis and favoring cell survival. GPX4 is differentially expressed according to myeloid differentiation stage, exhibiting lower expression in hematopoietic stem cells and polymorphonuclear leucocytes, while harboring higher level of expression in common myeloid progenitors and monocytes. In addition, GPX4 is highly expressed in most of acute myeloid leukemia (AML) subtypes compared to normal hematopoietic stem cells. High GPX4 expression is consistently correlated to poor prognosis in patients suffering AML. However, the role of GPX4 in the development of the myeloid lineage and in the initiation and progression of myeloid leukemia remains poorly explored. Given its essential role in the detoxification of lipid hydroperoxides, and its overexpression in most of myeloid malignancies, GPX4 inhibition has emerged as a promising therapeutic strategy to specifically trigger ferroptosis and eradicate myeloid leukemia cells. In this review, we describe the most recent advances concerning the role of GPX4 and, more generally ferroptosis in the myeloid lineage and in the emergence of AML. We also discuss the therapeutic interest and limitations of GPX4 inhibition alone or in combination with other drugs as innovative therapies to treat AML patients.

Keywords:  AML; Ferroptosis; GPX4; Myeloid differentiation; Myeloid leukemia; Small molecule inhibitors; Therapeutic strategies

DOI:  https://doi.org/10.1186/s11658-024-00613-6
Blood. 2024 Jul 11. 144(2): 126-128

CDK6: HSC fate and cell cycle nexus?

Dirk Loeffler.

DOI: https://doi.org/10.1182/blood.2024025085
Cytotherapy. 2024 Jun 06. pii: S1465-3249(24)00735-7. [Epub ahead of print]

International Society for Cell & Gene Therapy Stem Cell Engineering Committee report on the current state of hematopoietic stem and progenitor cell-based genomic therapies and the challenges faced.

Ashish O Gupta, Melissa Azul, Senthil Velan Bhoopalan, Allistair Abraham, Alice Bertaina, Alan Bidgoli, Carmem Bonfim, Amy DeZern, Jingjing Li, Chrystal U Louis, Duncan Purtill, Annalisa Ruggeri, Jaap Jan Boelens, Susan Prockop, Akshay Sharma.

  Genetic manipulation of hematopoietic stem cells (HSCs) is being developed as a therapeutic strategy for several inherited disorders. This field is rapidly evolving with several novel tools and techniques being employed to achieve desired genetic changes. While commercial products are now available for sickle cell disease, transfusion-dependent β-thalassemia, metachromatic leukodystrophy and adrenoleukodystrophy, several challenges remain in patient selection, HSC mobilization and collection, genetic manipulation of stem cells, conditioning, hematologic recovery and post-transplant complications, financial issues, equity of access and institutional and global preparedness. In this report, we explore the current state of development of these therapies and provide a comprehensive assessment of the challenges these therapies face as well as potential solutions.

Keywords:  adrenoleukodystrophy; gene editing; gene therapy; hematopoietic stem cells; metachromatic leukodystrophy; sickle cell disease; thalassemia

DOI:  https://doi.org/10.1016/j.jcyt.2024.06.002
Blood Adv. 2024 Jul 12. pii: bloodadvances.2024012867. [Epub ahead of print]

Differential inflammatory conditioning of the bone marrow by acute myeloid leukemia and its impact on progression.

Valentina R Minciacchi, Christina Karantanou, Jimena Bravo, Raquel S Pereira, Costanza Zanetti, Theresa Krack, Rahul Kumar, Katrin Bankov, Sylvia Hartmann, Brian Jp Huntly, Eshwar Meduri, Wolfram Ruf, Daniela S Krause.

Inflammation promotes solid tumor progression, but how regulatory mechanisms of inflammation may impact leukemia is less well studied. Using annexin A5 (ANXA 5), a calcium-binding protein known for apoptosis, which we discovered to be differentially expressed in the bone marrow microenvironment (BMM) of mice with acute myeloid (AML) versus chronic myeloid leukemia, as a model system, we unravel here a circuit in which AML-derived tumor necrosis factor (TNF)α dose-dependently reduces ANXA5 in the BMM. This creates an inflammatory BMM via elevated levels of prostaglandin E2 (PGE2). Via binding to its EP4 receptor, PGE2 increases -catenin and hypoxia-inducible factor (HIF) 1 α signaling in AML cells, thereby accelerating PGE2-sensitive AML. Human trephine biopsies may show lower ANXA5 expression and higher PGE2 expression in AML compared to other hematological malignancies. Further, syngeneic and xenogeneic transplantation models suggest a survival benefit after treatment with the inhibitor of prostaglandin-endoperoxide synthase 2 (cyclooxygenase 2 (COX2)), celecoxib, plus cytarabine in those AML types highly sensitive to PGE2 compared to cytarabine alone. Taken together, TNFα/ANXA5/NF-kB/COX2/PGE2-mediated inflammation influences AML course in a highly differential and circular manner, and AML patients with 'inflammatory AML' may benefit from antiphlogistic agents as adjunct therapy.

DOI: https://doi.org/10.1182/bloodadvances.2024012867
Nat Commun. 2024 Jul 07. 15(1): 5689

Hematopoietic stem and progenitor cell membrane-coated vesicles for bone marrow-targeted leukaemia drug delivery.

Jinxin Li, Honghui Wu, Zebin Yu, Qiwei Wang, Xin Zeng, Wenchang Qian, Siqi Lu, Lingli Jiang, Jingyi Li, Meng Zhu, Yingli Han, Jianqing Gao, Pengxu Qian.

Leukemia is a kind of hematological malignancy originating from bone marrow, which provides essential signals for initiation, progression, and recurrence of leukemia. However, how to specifically deliver drugs to the bone marrow remains elusive. Here, we develop biomimetic vesicles by infusing hematopoietic stem and progenitor cell (HSPC) membrane with liposomes (HSPC liposomes), which migrate to the bone marrow of leukemic mice via hyaluronic acid-CD44 axis. Moreover, the biomimetic vesicles exhibit superior binding affinity to leukemia cells through intercellular cell adhesion molecule-1 (ICAM-1)/integrin β2 (ITGB2) interaction. Further experiments validate that the vesicles carrying chemotherapy drug cytarabine (Ara-C@HSPC-Lipo) markedly inhibit proliferation, induce apoptosis and differentiation of leukemia cells, and decrease number of leukemia stem cells. Mechanically, RNA-seq reveals that Ara-C@HSPC-Lipo treatment induces apoptosis and differentiation and inhibits the oncogenic pathways. Finally, we verify that HSPC liposomes are safe in mice. This study provides a method for targeting bone marrow and treating leukemia.

DOI: https://doi.org/10.1038/s41467-024-50021-9
J Proteome Res. 2024 Jul 09.

Are We There Yet? Assessing the Readiness of Single-Cell Proteomics to Answer Biological Hypotheses.

Alyssa A Nitz, Jose Humberto Giraldez Chavez, Zachary G Eliason, Samuel H Payne.

  Single-cell analysis is an active area of research in many fields of biology. Measurements at single-cell resolution allow researchers to study diverse populations without losing biologically meaningful information to sample averages. Many technologies have been used to study single cells, including mass spectrometry-based single-cell proteomics (SCP). SCP has seen a lot of growth over the past couple of years through improvements in data acquisition and analysis, leading to greater proteomic depth. Because method development has been the main focus in SCP, biological applications have been sprinkled in only as proof-of-concept. However, SCP methods now provide significant coverage of the proteome and have been implemented in many laboratories. Thus, a primary question to address in our community is whether the current state of technology is ready for widespread adoption for biological inquiry. In this Perspective, we examine the potential for SCP in three thematic areas of biological investigation: cell annotation, developmental trajectories, and spatial mapping. We identify that the primary limitation of SCP is sample throughput. As proteome depth has been the primary target for method development to date, we advocate for a change in focus to facilitate measuring tens of thousands of single-cell proteomes to enable biological applications beyond proof-of-concept.

Keywords:  algorithms; experimental design; single-cell proteomics; statistical rigor

DOI:  https://doi.org/10.1021/acs.jproteome.4c00091
Circ Res. 2024 Jul 11.

RNF149 Destabilizes IFNGR1 in Macrophages to Favor Post-Infarction Cardiac Repair.

Chun-Kai Huang, Zhiyong Chen, Zhongxing Zhou, Shuaijie Chen, Longqing Chen, Liliang Li, Tao Li, Xiaoxiang Yan, Dajun Chai.

   BACKGROUND: Macrophage-driven inflammation critically involves in cardiac injury and repair following myocardial infarction (MI). However, the intrinsic mechanisms that halt the immune response of macrophages, which is critical to preserve homeostasis and effective infarct repair, remain to be fully defined. Here, we aimed to determine the ubiquitination-mediated regulatory effects on averting exaggerated inflammatory responses in cardiac macrophages.
METHODS: We used transcriptome analysis of mouse cardiac macrophages and bone marrow-derived macrophages to identify the E3 ubiquitin ligase RNF149 (RING finger protein 149) as a modulator of macrophage response to MI. Employing loss-of-function methodologies, bone marrow transplantation approaches, and adenovirus-mediated RNF149 overexpression in macrophages, we elucidated the functional role of RNF149 in MI. We explored the underlying mechanisms through flow cytometry, transcriptome analysis, immunoprecipitation/mass spectrometry analysis, and functional experiments. RNF149 expression was measured in the cardiac tissues of patients with acute MI and healthy controls.
RESULTS: RNF149 was highly expressed in murine and human cardiac macrophages at the early phase of MI. Knockout of RNF149, transplantation of Rnf149-/- bone marrow, and bone marrow macrophage-specific RNF149-knockdown markedly exacerbated cardiac dysfunction in murine MI models. Conversely, overexpression of RNF149 in macrophages attenuated the ischemia-induced decline in cardiac contractile function. RNF149 deletion increased infiltration of proinflammatory monocytes/macrophages, accompanied by a hastened decline in reparative subsets, leading to aggravation of myocardial apoptosis and impairment of infarct healing. Our data revealed that RNF149 in infiltrated macrophages restricted inflammation by promoting ubiquitylation-dependent proteasomal degradation of IFNGR1 (interferon gamma receptor 1). Loss of IFNGR1 rescued deleterious effects of RNF149 deficiency on MI. We further demonstrated that STAT1 activation induced Rnf149 transcription, which, in turn, destabilized the IFNGR1 protein to counteract type-II IFN (interferon) signaling, creating a feedback control mechanism to fine-tune macrophage-driven inflammation.
CONCLUSIONS: These findings highlight the significance of RNF149 as a molecular brake on macrophage response to MI and uncover a macrophage-intrinsic posttranslational mechanism essential for maintaining immune homeostasis and facilitating cardiac repair following MI.

Keywords:  cardiovascular diseases; inflammation; macrophages; myocardial infarction; ubiquitination

DOI:  https://doi.org/10.1161/CIRCRESAHA.123.324023
Cancer Lett. 2024 Jul 08. pii: S0304-3835(24)00503-2. [Epub ahead of print] 217108

Targeting of human cancer stem cells predicts efficacy and toxicity of FDA-approved oncology drugs.

Kinga Vojnits, Zhuohang Feng, Paige Johnson, Sibylle Pfammatter, Pierre Thibault, Mick Bhatia.

  Cancer remains the leading cause of death worldwide with approved oncology drugs continuing to have heterogenous patient responses and accompanied adverse effects (AEs) that limits effectiveness. Here, we examined >100 FDA-approved oncology drugs in the context of stemness using a surrogate model of transformed human pluripotent cancer stem cells (CSCs) vs. healthy stem cells (hSCs) capable of distinguishing abnormal self-renewal and differentiation. Although a proportion of these drugs had no effects (inactive), a larger portion affected CSCs (active), and a unique subset preferentially affected CSCs over hSCs (selective). Single cell gene expression and protein profiling of each drug's FDA recognized target provided a molecular correlation of responses in CSCs vs. hSCs. Uniquely, drugs selective for CSCs demonstrated clinical efficacy, measured by overall survival, and reduced AEs. Our findings reveal that while unintentional, half of anticancer drugs are active against CSCs and associated with improved clinical outcomes. Based on these findings, we suggest ability to target CSC targeting should be included as a property of early onco-therapeutic development.

Keywords:  drug selectivity; efficacy; oncology drugs; phenotypic screen; scRNAseq; stemness

DOI:  https://doi.org/10.1016/j.canlet.2024.217108
Heliyon. 2024 Jun 30. 10(12): e32974

Aging-related changes in the mechanical properties of single cells.

Amarnath Singam, Chandrabali Bhattacharya, Seungman Park.

  Mechanical properties, along with biochemical and molecular properties, play crucial roles in governing cellular function and homeostasis. Cellular mechanics are influenced by various factors, including physiological and pathological states, making them potential biomarkers for diseases and aging. While several methods such as AFM, particle-tracking microrheology, optical tweezers/stretching, magnetic tweezers/twisting cytometry, microfluidics, and micropipette aspiration have been widely utilized to measure the mechanical properties of single cells, our understanding of how aging affects these properties remains limited. To fill this knowledge gap, we provide a brief overview of the commonly used methods to measure single-cell mechanical properties. We then delve into the effects of aging on the mechanical properties of different cell types. Finally, we discuss the importance of studying cellular viscous and viscoelastic properties as well as aging induced by different stressors to gain a deeper understanding of the aging process and aging-related diseases.

Keywords:  Aging; Cell; Cytoskeleton; Elastic property; F-actin; Mechanical property; Stress fiber; Viscous property

DOI:  https://doi.org/10.1016/j.heliyon.2024.e32974
Life Sci Alliance. 2024 Sep;pii: e202402765. [Epub ahead of print]7(9):

Dual-localized PPTC7 limits mitophagy through proximal and dynamic interactions with BNIP3 and NIX.

Lianjie Wei, Mehmet Oguz Gok, Jordyn D Svoboda, Keri-Lyn Kozul, Merima Forny, Jonathan R Friedman, Natalie M Niemi.

PPTC7 is a mitochondrial-localized phosphatase that suppresses BNIP3- and NIX-mediated mitophagy, but the mechanisms underlying this regulation remain ill-defined. Here, we demonstrate that loss of PPTC7 upregulates BNIP3 and NIX post-transcriptionally and independent of HIF-1α stabilization. Loss of PPTC7 prolongs the half-life of BNIP3 and NIX while blunting their accumulation in response to proteasomal inhibition, suggesting that PPTC7 promotes the ubiquitin-mediated turnover of BNIP3 and NIX. Consistently, overexpression of PPTC7 limits the accumulation of BNIP3 and NIX protein levels, which requires an intact catalytic motif but is surprisingly independent of its targeting to mitochondria. Consistently, we find that PPTC7 is dual-localized to the outer mitochondrial membrane and the matrix. Importantly, anchoring PPTC7 to the outer mitochondrial membrane is sufficient to blunt BNIP3 and NIX accumulation, and proximity labeling and fluorescence co-localization experiments demonstrate that PPTC7 dynamically associates with BNIP3 and NIX within the native cellular environment. Collectively, these data reveal that a fraction of PPTC7 localizes to the outer mitochondrial membrane to promote the proteasomal turnover of BNIP3 and NIX, limiting basal mitophagy.

DOI: https://doi.org/10.26508/lsa.202402765
J Cell Physiol. 2024 Jul 10. e31371

Functional characterization of native Piezo1 as calcium and magnesium influx pathway in human myeloid leukemia cells.

Valeria Y Vasileva, Daria V Lysikova, Anastasia V Sudarikova, Zuleikha M Khairullina, Polina I Kirillova, Elena A Morachevskaya, Vladislav I Chubinskiy-Nadezhdin.

  Piezo1 is a Ca2+-permeable mechanically activated ion channel that is involved in various physiological processes and cellular responses to mechanical stimuli. The study of biophysical characteristics of Piezo1 is important for understanding the mechanisms of its function and regulation. Stretch activation, a routine approach that is applied to stimulate Piezo1 activity in the plasma membrane, has a number of significant limitations that complicate precise single-channel analysis. Here, we aimed to determine pore properties of native Piezo1, specifically to examine permeation for physiologically relevant signaling divalent ions (calcium and magnesium) in human myeloid leukemia K562 cells using Piezo1-specific chemical agonist, Yoda1. Using a combination of low-noise single-current patch-clamp recordings of Piezo1 activity in response to Yoda1, we have determined single-channel characteristics of native Piezo1 under various ionic conditions. Whole-cell assay allowed us to directly measure Piezo1 single currents carried by Ca2+ or Mg2+ ions in the absence of other permeable cations in the extracellular solutions; unitary conductance values estimated at various concentrations of Mg2+ revealed strong saturation effect. Patch clamp data complemented with fluorescent imaging clearly evidenced Ca2+ and Mg2+ entry via native Piezo1 channel in human leukemia K562 cells. Mg2+ influx via Piezo1 was detected under quasi-physiological conditions, thus showing that Piezo1 channels could potentially provide the physiological relevant pathway for Mg2+ ion transport and contribute to the regulation of Mg2+-dependent intracellular signaling.

Keywords:  K562 cells; Piezo1; Yoda1; calcium; magnesium; single‐channel analysis

DOI:  https://doi.org/10.1002/jcp.31371
Cell Prolif. 2024 Jul 09. e13715

Heterogeneous stiffness of the bone marrow microenvironment regulates the fate decision of haematopoietic stem and progenitor cells.

Guolin Shi, Zhuo Chang, Pan Zhang, Xiaohang Zou, Xinmin Zheng, Xiru Liu, Jinxiao Yan, Huiyun Xu, Zhenhao Tian, Nu Zhang, Ning Cui, Leming Sun, Guangkui Xu, Hui Yang.

The bone marrow (BM) niches are the complex microenvironments that surround cells, providing various external stimuli to regulate a range of haematopoietic stem cell (HSC) behaviours. Recently, it has been proposed that the fate decision of HSCs is often correlated with significantly altered biophysical signals of BM niches. To thoroughly elucidate the effect of mechanical microenvironments on cell fates, we constructed 2D and 3D cell culture hydrogels using polyacrylamide to replicate the mechanical properties of heterogeneous sub-niches, including the inherent rigidity of marrow adipose tissue (2 kPa), perivascular tissue (8 kPa) and endosteum region (35 kPa) in BM. Our observations suggest that HSCs can respond to the mechanical heterogeneity of the BM microenvironment, exhibiting diversity in cell mechanics, haematopoietic pool maintenance and differentiated lineages. Hydrogels with higher stiffness promote the preservation of long-term repopulating HSCs (LT-HSCs), while those with lower stiffness support multi-potent progenitors (MPPs) viability in vitro. Furthermore, we established a comprehensive transcriptional profile of haematopoietic subpopulations to reflect the multipotency of haematopoietic stem and progenitor cells (HSPCs) that are modulated by niche-like stiffness. Our findings demonstrate that HSPCs exhibit completely distinct downstream differentiated preferences within hydrogel systems of varying stiffness. This highlights the crucial role of tissue-specific mechanical properties in HSC lineage decisions, which may provide innovative solutions to clinical challenges.

DOI: https://doi.org/10.1111/cpr.13715
Nat Commun. 2024 Jul 06. 15(1): 5678

Erythroid-intrinsic activation of TLR8 impairs erythropoiesis in inherited anemia.

Jing Liang, Yang Wan, Jie Gao, Lingyue Zheng, Jingwei Wang, Peng Wu, Yue Li, Bingrui Wang, Ding Wang, Yige Ma, Biao Shen, Xue Lv, Di Wang, Na An, Xiaoli Ma, Guangfeng Geng, Jingyuan Tong, Jinhua Liu, Guo Chen, Meng Gao, Ryo Kurita, Yukio Nakamura, Ping Zhu, Hang Yin, Xiaofan Zhu, Lihong Shi.

Inherited non-hemolytic anemia is a group of rare bone marrow disorders characterized by erythroid defects. Although concerted efforts have been made to explore the underlying pathogenetic mechanisms of these diseases, the understanding of the causative mutations are still incomplete. Here we identify in a diseased pedigree that a gain-of-function mutation in toll-like receptor 8 (TLR8) is implicated in inherited non-hemolytic anemia. TLR8 is expressed in erythroid lineage and erythropoiesis is impaired by TLR8 activation whereas enhanced by TLR8 inhibition from erythroid progenitor stage. Mechanistically, TLR8 activation blocks annexin A2 (ANXA2)-mediated plasma membrane localization of STAT5 and disrupts EPO signaling in HuDEP2 cells. TLR8 inhibition improves erythropoiesis in RPS19+/- HuDEP2 cells and CD34+ cells from healthy donors and inherited non-hemolytic anemic patients. Collectively, we identify a gene implicated in inherited anemia and a previously undescribed role for TLR8 in erythropoiesis, which could potentially be explored for therapeutic benefit in inherited anemia.

DOI: https://doi.org/10.1038/s41467-024-50066-w
Nature. 2024 Jul 10.

Plasmacytoid dendritic cells control homeostasis of megakaryopoiesis.

Florian Gaertner, Hellen Ishikawa-Ankerhold, Susanne Stutte, Wenwen Fu, Jutta Weitz, Anne Dueck, Bhavishya Nelakuditi, Valeria Fumagalli, Dominic van den Heuvel, Larissa Belz, Gulnoza Sobirova, Zhe Zhang, Anna Titova, Alejandro Martinez Navarro, Kami Pekayvaz, Michael Lorenz, Louisa von Baumgarten, Jan Kranich, Tobias Straub, Bastian Popper, Vanessa Zheden, Walter Anton Kaufmann, Chenglong Guo, Guido Piontek, Saskia von Stillfried, Peter Boor, Marco Colonna, Sebastian Clauß, Christian Schulz, Thomas Brocker, Barbara Walzog, Christoph Scheiermann, William C Aird, Claus Nerlov, Konstantin Stark, Tobias Petzold, Stefan Engelhardt, Michael Sixt, Robert Hauschild, Martina Rudelius, Robert A J Oostendorp, Matteo Iannacone, Matthias Heinig, Steffen Massberg.

Platelet homeostasis is essential for vascular integrity and immune defence1,2. Although the process of platelet formation by fragmenting megakaryocytes (MKs; thrombopoiesis) has been extensively studied, the cellular and molecular mechanisms required to constantly replenish the pool of MKs by their progenitor cells (megakaryopoiesis) remains unclear3,4. Here we use intravital imaging to track the cellular dynamics of megakaryopoiesis over days. We identify plasmacytoid dendritic cells (pDCs) as homeostatic sensors that monitor the bone marrow for apoptotic MKs and deliver IFNα to the MK niche triggering local on-demand proliferation and maturation of MK progenitors. This pDC-dependent feedback loop is crucial for MK and platelet homeostasis at steady state and under stress. pDCs are best known for their ability to function as vigilant detectors of viral infection5. We show that virus-induced activation of pDCs interferes with their function as homeostatic sensors of megakaryopoiesis. Consequently, activation of pDCs by SARS-CoV-2 leads to excessive megakaryopoiesis. Together, we identify a pDC-dependent homeostatic circuit that involves innate immune sensing and demand-adapted release of inflammatory mediators to maintain homeostasis of the megakaryocytic lineage.

DOI: https://doi.org/10.1038/s41586-024-07671-y
Trends Cancer. 2024 Jul 05. pii: S2405-8033(24)00116-X. [Epub ahead of print]

Targeting cancer stem cells in multiple myeloma.

Emine Gulsen Gunes, Metin Gunes, Jianhua Yu, Murali Janakiram.

  Multiple myeloma (MM) is a hematological malignancy of bone marrow (BM) plasma cells with excessive clonal expansion and is associated with the overproduction of light-chain or monoclonal immunoglobulins (Igs). MM remains incurable, with high rates of relapses and refractory disease after first-line treatment. Cancer stem cells (CSCs) have been implicated in drug resistance in MM; however, the evidence for CSCs in MM is not adequate, partly due to a lack of uniformity in the definitions of multiple myeloma stem cells (MMSCs). We review advances in understanding MMSCs and their role in drug resistance to MM therapies. We also discuss novel therapeutic strategies to overcome MMSC-mediated relapses and drug resistance.

Keywords:  cancer stem cells; drug resistance; multiple myeloma; refractory; relapse

DOI:  https://doi.org/10.1016/j.trecan.2024.06.002
Biomaterials. 2024 Jun 26. pii: S0142-9612(24)00218-7. [Epub ahead of print]311 122684

A 3D micro-printed single cell micro-niche with asymmetric niche signals - An in vitro model for asymmetric cell division study.

Nan Huang, Barbara Pui Chan.

  Intricate microenvironment signals orchestrate to affect cell behavior and fate during tissue morphogenesis. However, the underlying mechanisms on how specific local niche signals influence cell behavior and fate are not fully understood, owing to the lack of in vitro platform able to precisely, quantitatively, spatially, and independently manipulate individual niche signals. Here, microarrays of protein-based 3D single cell micro-niche (3D-SCμN), with precisely engineered biophysical and biochemical niche signals, are micro-printed by a multiphoton microfabrication and micropatterning technology. Mouse embryonic stem cell (mESC) is used as the model cell to study how local niche signals affect stem cell behavior and fate. By precisely engineering the internal microstructures of the 3D SCμNs, we demonstrate that the cell division direction can be controlled by the biophysical niche signals, in a cell shape-independent manner. After confining the cell division direction to a dominating axis, single mESCs are exposed to asymmetric biochemical niche signals, specifically, cell-cell adhesion molecule on one side and extracellular matrix on the other side. We demonstrate that, symmetry-breaking (asymmetric) niche signals successfully trigger cell polarity formation and bias the orientation of asymmetric cell division, the mitosis process resulting in two daughter cells with differential fates, in mESCs.

Keywords:  3D single cell micro-niche; Asymmetric cell division; Cell fate; Cell-cell interaction; Cell-matrix interaction; Mouse embryonic stem cell; Multiphoton microfabrication

DOI:  https://doi.org/10.1016/j.biomaterials.2024.122684
Nat Commun. 2024 Jul 07. 15(1): 5693

Epigenetic alterations affecting hematopoietic regulatory networks as drivers of mixed myeloid/lymphoid leukemia.

Roger Mulet-Lazaro, Stanley van Herk, Margit Nuetzel, Aniko Sijs-Szabo, Noelia Díaz, Katherine Kelly, Claudia Erpelinck-Verschueren, Lucia Schwarzfischer-Pfeilschifter, Hanna Stanewsky, Ute Ackermann, Dagmar Glatz, Johanna Raithel, Alexander Fischer, Sandra Pohl, Anita Rijneveld, Juan M Vaquerizas, Christian Thiede, Christoph Plass, Bas J Wouters, Ruud Delwel, Michael Rehli, Claudia Gebhard.

Leukemias with ambiguous lineage comprise several loosely defined entities, often without a clear mechanistic basis. Here, we extensively profile the epigenome and transcriptome of a subgroup of such leukemias with CpG Island Methylator Phenotype. These leukemias exhibit comparable hybrid myeloid/lymphoid epigenetic landscapes, yet heterogeneous genetic alterations, suggesting they are defined by their shared epigenetic profile rather than common genetic lesions. Gene expression enrichment reveals similarity with early T-cell precursor acute lymphoblastic leukemia and a lymphoid progenitor cell of origin. In line with this, integration of differential DNA methylation and gene expression shows widespread silencing of myeloid transcription factors. Moreover, binding sites for hematopoietic transcription factors, including CEBPA, SPI1 and LEF1, are uniquely inaccessible in these leukemias. Hypermethylation also results in loss of CTCF binding, accompanied by changes in chromatin interactions involving key transcription factors. In conclusion, epigenetic dysregulation, and not genetic lesions, explains the mixed phenotype of this group of leukemias with ambiguous lineage. The data collected here constitute a useful and comprehensive epigenomic reference for subsequent studies of acute myeloid leukemias, T-cell acute lymphoblastic leukemias and mixed-phenotype leukemias.

DOI: https://doi.org/10.1038/s41467-024-49811-y
Proc Natl Acad Sci U S A. 2024 Jul 16. 121(29): e2321647121

FAM110A promotes mitotic spindle formation by linking microtubules with actin cytoskeleton.

Cecilia Aquino-Perez, Mahira Safaralizade, Roman Podhajecky, Hong Wang, Zdenek Lansky, Robert Grosse, Libor Macurek.

  Precise segregation of chromosomes during mitosis requires assembly of a bipolar mitotic spindle followed by correct attachment of microtubules to the kinetochores. This highly spatiotemporally organized process is controlled by various mitotic kinases and molecular motors. We have recently shown that Casein Kinase 1 (CK1) promotes timely progression through mitosis by phosphorylating FAM110A leading to its enrichment at spindle poles. However, the mechanism by which FAM110A exerts its function in mitosis is unknown. Using structure prediction and a set of deletion mutants, we mapped here the interaction of the N- and C-terminal domains of FAM110A with actin and tubulin, respectively. Next, we found that the FAM110A-Δ40-61 mutant deficient in actin binding failed to rescue defects in chromosomal alignment caused by depletion of endogenous FAM110A. Depletion of FAM110A impaired assembly of F-actin in the proximity of spindle poles and was rescued by expression of the wild-type FAM110A, but not the FAM110A-Δ40-61 mutant. Purified FAM110A promoted binding of F-actin to microtubules as well as bundling of actin filaments in vitro. Finally, we found that the inhibition of CK1 impaired spindle actin formation and delayed progression through mitosis. We propose that CK1 and FAM110A promote timely progression through mitosis by mediating the interaction between spindle microtubules and filamentous actin to ensure proper mitotic spindle formation.

Keywords:  actin; microtubules; mitosis; mitotic spindle; protein kinase

DOI:  https://doi.org/10.1073/pnas.2321647121
Nat Chem Biol. 2024 Jul 08.

Distinct phases of cellular signaling revealed by time-resolved protein synthesis.

Gihoon Lee, Tom W Muir.

The post-translational regulation of protein function is involved in most cellular processes. As such, synthetic biology tools that operate at this level provide opportunities for manipulating cellular states. Here we deploy proximity-triggered protein trans-splicing technology to enable the time-resolved synthesis of target proteins from premade parts. The modularity of the strategy allows for the addition or removal of various control elements as a function of the splicing reaction, in the process permitting the cellular location and/or activity state of starting materials and products to be differentiated. The approach is applied to a diverse set of proteins, including the kinase oncofusions breakpoint cluster region-Abelson (BCR-ABL) and DNAJ-PKAc where dynamic cellular phosphorylation events are dissected, revealing distinct phases of signaling and identifying molecular players connecting the oncofusion to cancer transformation as new therapeutic targets of cancer cells. We envision that the tools and control strategies developed herein will allow the activity of both naturally occurring and designer proteins to be harnessed for basic and applied research.

DOI: https://doi.org/10.1038/s41589-024-01677-3
Proc Natl Acad Sci U S A. 2024 Jul 16. 121(29): e2320769121

Cell-intrinsic mechanical regulation of plasma membrane accumulation at the cytokinetic furrow.

Roberto Alonso-Matilla, Alice R Lam, Teemu P Miettinen.

  Cytokinesis is the process where the mother cell's cytoplasm separates into daughter cells. This is driven by an actomyosin contractile ring that produces cortical contractility and drives cleavage furrow ingression, resulting in the formation of a thin intercellular bridge. While cytoskeletal reorganization during cytokinesis has been extensively studied, less is known about the spatiotemporal dynamics of the plasma membrane. Here, we image and model plasma membrane lipid and protein dynamics on the cell surface during leukemia cell cytokinesis. We reveal an extensive accumulation and folding of the plasma membrane at the cleavage furrow and the intercellular bridge, accompanied by a depletion and unfolding of the plasma membrane at the cell poles. These membrane dynamics are caused by two actomyosin-driven biophysical mechanisms: the radial constriction of the cleavage furrow causes local compression of the apparent cell surface area and accumulation of the plasma membrane at the furrow, while actomyosin cortical flows drag the plasma membrane toward the cell division plane as the furrow ingresses. The magnitude of these effects depends on the plasma membrane fluidity, cortex adhesion, and cortical contractility. Overall, our work reveals cell-intrinsic mechanical regulation of plasma membrane accumulation at the cleavage furrow that is likely to generate localized differences in membrane tension across the cytokinetic cell. This may locally alter endocytosis, exocytosis, and mechanotransduction, while also serving as a self-protecting mechanism against cytokinesis failures that arise from high membrane tension at the intercellular bridge.

Keywords:  actomyosin; biophysical model; cytokinesis; membrane tension; plasma membrane

DOI:  https://doi.org/10.1073/pnas.2320769121