Tracking of Normal and Malignant Progenitor Cell Cycle Transit in a Defined Niche

Pineda, Gabriel; Lennon, Kathleen M.; Santos, Nathaniel Delos; Lambert-Fliszar, Florence; Riso, Gennarina L.; Lazzari, Elisa; Marra, Marco A.; Morris, Sheldon R.; Sakaue-Sawano, Asako; Miyawaki, Atsushi

doi:10.1038/srep23885

Cited by 12 publications

(18 citation statements)

References 36 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…To better quantify cell cycle progression in single human MSC from young and aged donors, we generated a lentiviral bicistronic reporter vector encoding fluorescent ubiquitination‐based cell cycle indicator probes (Fucci system). The lentiviral vector expresses mVenus‐hGeminin(1/110) fused to mCherry‐hCdt1(30/120) by the T2A peptide using an EF1α promoter that generates optimal levels of gene expression in primary cells (Pineda et al, ). These fluorescent reporters allow us to discriminate between three cell cycle phases as G1 by red fluorescence, G1/S by yellow fluorescence, and S/G2/M by green fluorescence.…”

Section: Resultsmentioning

confidence: 99%

An early‐senescence state in aged mesenchymal stromal cells contributes to hematopoietic stem and progenitor cell clonogenic impairment through the activation of a pro‐inflammatory program

et al. 2019

View full text Add to dashboard Cite

Hematopoietic stem and progenitor cells (HSPC) reside in the bone marrow (BM) niche and serve as a reservoir for mature blood cells throughout life. Aging in the BM is characterized by low‐grade chronic inflammation that could contribute to the reduced functionality of aged HSPC. Mesenchymal stromal cells (MSC) in the BM support HSPC self‐renewal. However, changes in MSC function with age and the crosstalk between MSC and HSPC remain understudied. Here, we conducted an extensive characterization of senescence features in BM‐derived MSC from young and aged healthy donors. Aged MSC displayed an enlarged senescent‐like morphology, a delayed clonogenic potential and reduced proliferation ability when compared to younger counterparts. Of note, the observed proliferation delay was associated with increased levels of SA‐β‐galactosidase (SA‐β‐Gal) and lipofuscin in aged MSC at early passages and a modest but consistent accumulation of physical DNA damage and DNA damage response (DDR) activation. Consistent with the establishment of a senescence‐like state in aged MSC, we detected an increase in pro‐inflammatory senescence‐associated secretory phenotype (SASP) factors, both at the transcript and protein levels. Conversely, the immunomodulatory properties of aged MSC were significantly reduced. Importantly, exposure of young HSPC to factors secreted by aged MSC induced pro‐inflammatory genes in HSPC and impaired HSPC clonogenic potential in a SASP‐dependent manner. Altogether, our results reveal that BM‐derived MSC from aged healthy donors display features of senescence and that, during aging, MSC‐associated secretomes contribute to activate an inflammatory transcriptional program in HSPC that may ultimately impair their functionality.

show abstract

Section: Resultsmentioning

confidence: 99%

An early‐senescence state in aged mesenchymal stromal cells contributes to hematopoietic stem and progenitor cell clonogenic impairment through the activation of a pro‐inflammatory program

et al. 2019

View full text Add to dashboard Cite

show abstract

“…Cell cycle state sensors have been used for the last decade in tissue culture and in vivo to provide new insights into underlying cell biology. Various cell cycle sensors have been developed, starting with FUCCI in 2008 (Sakaue-Sawano et al, 2008), each optimized for different research paradigms (Abe et al, 2013;Bouldin and Kimelman, 2014b;Fukuhara et al, 2014;Ogura et al, 2011;Pineda et al, 2016;Ridenour et al, 2012;Sakaue-Sawano et al, 2017;Sugiyama et al, 2009;Zielke et al, 2016). While FUCCI-style biosensors have been developed to better assess G0/G1 in zebrafish using inactivated p27 (Oki et al, 2014), and in tissue culture by assessing the primary cilium cycle (Ford et al, 2018), they require multiple transgenes to readout cell cycle states accurately.…”

Section: Discussionmentioning

confidence: 99%

Visualizing the metazoan proliferation-terminal differentiation decisionin vivo

Kohrman

Adikes

Martinez

et al. 2019

Preprint

View full text Add to dashboard Cite

During organismal development, differential regulation of the cell cycle is critical to many cell biological processes, including cell fate specification and differentiation. While the mechanisms of cell cycle regulation are well studied, how control of the cell cycle is linked to differentiated cellular behavior remains poorly understood, mostly due to our inability to directly and precisely measure cell cycle state. In order to characterize cell cycle state live, we adapted a cyclindependent kinase (CDK) biosensor for in vivo use in the roundworm nematode, Caenorhabditis elegans. The CDK biosensor measures the cytoplasmic-to-nuclear localization of a portion of human DNA Helicase B (DHB) fused to a fluorescent protein to assess cell cycle state. The dynamic localization of DHB results from phosphorylation of the biosensor by CDKs, thereby allowing for quantitative assessment of cell cycle state. We demonstrate here the use of this biosensor to quantify lineage-specific differences between cycling cells and to examine the proliferation-differentiation decision. Unlike other live cell imaging tools (e.g., FUCCI), we show that DHB can be used to distinguish between actively cycling cells in the G1 phase of the cell cycle and terminally differentiated cells exited in G0. Thus, we provide here a new resource to study the control and timing of the metazoan cell cycle during cell fate specification and differentiation.

show abstract

“…An EcoRI-containing double-stranded synthetic oligonucleotide spanning the −109 bp MIR300 regulatory (region and containing the T/G and C/G mutated C/EBPβ consensus binding sites located at positions -64 and -46, respectively, was subcloned into EcoRI-digested pGF1 vector. pCDH-Flag-SET, fluorescent ubiquitination-based cell-cycle indicator reporter pCDH-FUCCI2BL, MigR1-ΔuORF-C/EBPβ-ER TAM , and MigR1-ΔuORF-C/EBPα-HA constructs were described previously (25,(62)(63)(64).…”

Section: P109-c/ebpbmut-gfp/lucmentioning

confidence: 99%

Persistence of Drug-Resistant Leukemic Stem Cells and Impaired NK Cell Immunity in CML Patients Depend on MIR300 Antiproliferative and PP2A-Activating Functions

Silvestri

Trotta

Stramucci

et al. 2020

Blood Cancer Discovery

Self Cite

View full text Add to dashboard Cite

Persistence of drug-resistant quiescent leukemic stem cells (LSC) and impaired natural killer (NK) cell immune response account for relapse of chronic myelogenous leukemia (CML). Inactivation of protein phosphatase 2A (PP2A) is essential for CML-quiescent LSC survival and NK cell antitumor activity. Here we show that MIR300 has antiproliferative and PP2A-activating functions that are dose dependently differentially induced by CCND2/CDK6 and SET inhibition, respectively. MIR300 is upregulated in CML LSCs and NK cells by bone marrow microenvironment (BMM) signals to induce quiescence and impair immune response, respectively. Conversely, BCR-ABL1 downregulates MIR300 in CML progenitors to prevent growth arrest and PP2A-mediated apoptosis. Quiescent LSCs escape apoptosis by upregulating TUG1 long noncoding RNA that uncouples and limits MIR300 function to cytostasis. Genetic and pharmacologic MIR300 modulation and/or PP2A-activating drug treatment restore NK cell activity, inhibit BMM-induced growth arrest, and selectively trigger LSC apoptosis in vitro and in patient-derived xenografts; hence, the importance of MIR300 and PP2A activity for CML development and therapy. SIGNIFICANCE:Tumor-naïve microenvironment-induced MIR300 is the only tumor suppressor miRNA that induces CML LSC quiescence while inhibiting NK cell antitumor immune response and CML LSC/ progenitor cell apoptosis through its anti-proliferative and PP2A-activating functions, respectively. Thus, the importance of MIR300 and PP2A-activating drugs for formation/survival and eradication of drug-resistant CML LSCs, respectively.

show abstract

Tracking of Normal and Malignant Progenitor Cell Cycle Transit in a Defined Niche

Cited by 12 publications

References 36 publications

An early‐senescence state in aged mesenchymal stromal cells contributes to hematopoietic stem and progenitor cell clonogenic impairment through the activation of a pro‐inflammatory program

An early‐senescence state in aged mesenchymal stromal cells contributes to hematopoietic stem and progenitor cell clonogenic impairment through the activation of a pro‐inflammatory program

Visualizing the metazoan proliferation-terminal differentiation decisionin vivo

Persistence of Drug-Resistant Leukemic Stem Cells and Impaired NK Cell Immunity in CML Patients Depend on MIR300 Antiproliferative and PP2A-Activating Functions

Contact Info

Product

Resources

About