Proliferation and Self-Renewal Are Differentially Sensitive to NRASG12V Oncogene Levels in an Acute Myeloid Leukemia Cell Line

Morito Kurata, Marie Lue Antony, Klara E. Noble-Orcutt, Susan K. Rathe, Yoonkyu Lee, Hidehiro Furuno, Sachiko Ishibashi, Masumi Ikeda, Kouhei Yamamoto, Masanobu Kitagawa, David A. Largaespada, Zohar Sachs

Research output: Contribution to journalArticlepeer-review

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NRAS proteins are central regulators of proliferation, survival, and self-renewal in leukemia. Previous work demonstrated that the effects of oncogenic NRAS in mediating proliferation and self-renewal are mutually exclusive within leukemia subpopulations and that levels of oncogenic NRAS vary between highly proliferative and self-renewing leukemia subpopulations. These findings suggest that NRAS activity levels may be important determinants of leukemic behavior. To define how oncogenic NRAS levels affect these functions, we genetically engineered an acute myeloid leukemia (AML) cell line, THP-1, to express variable levels of NRASG12V. We replaced the endogenous NRASG12D gene with a tetracycline-inducible and dose-responsive NRASG12V transgene. Cells lacking NRASG12V oncoprotein were cell-cycle arrested. Intermediate levels of NRASG12V induced maximal proliferation; higher levels led to attenuated proliferation, increased G1 arrest, senescence markers, and maximal self-renewal capacity. Higher levels of the oncoprotein also induced self-renewal and mitochondrial genes. We used mass cytometry (CyTOF) to define the downstream signaling events that mediate these differential effects. Not surprisingly, we found that the levels of such canonical RAS-effectors as pERK and p4EBP1 correlated with NRASG12V levels. b-Catenin, a mediator of self-renewal, also correlated with NRASG12V levels. These signaling intermediates may mediate the differential effects of NRASG12V in leukemia biology. Together, these data reveal that oncogenic NRAS levels are important determinants of leukemic behavior explaining heterogeneity in phenotypes within a clone. This system provides a new model to study RAS oncogene addiction and RAS-induced self-renewal in AML.

Original languageEnglish (US)
Pages (from-to)1646-1658
Number of pages13
JournalMolecular Cancer Research
Issue number11
StatePublished - Nov 2022

Bibliographical note

Funding Information:
Dr. Kyle Williams and Mr. Rory Williams helped the analysis of active Ras Pull-Down assay. Funding for this project was provided by The Leukemia & Lymphoma Society (grant no. 7019-04), Grant-in-Aid for Scientific Research (C; 18K06953) from the Japan Society for the Promotion of Science, and Kawano Masanori Memorial Foundation for Promotion of Pediatrics. Z. Sachs was supported by the American Cancer Society, Frederick A. DeLuca Foundation, Mentored Research Scholar Grant (MRSG-16-195-01-DDC); the Lois and Richard King Assistant Professorship in Medicine at the University of Minnesota, the Clinical and Translational Science Institute at the University of Minnesota KL2 Career Development Award and K to R01 Award (NIH/NCATS ULI RR033183 and KL2 RR0333182); the American Cancer Society Institutional Research Grant at the University of Minnesota (124166-IRG-58-001-55-IRG12); the Masonic Cancer Center at the University of Minnesota Translational Working Group Award and Genetic Mechanisms of Cancer Award; the University of Minnesota Department of Medicine Women’s Early Research Career Award; the division of Hematology, Oncology, and Transplantation, Department of Medicine; and the University of Minnesota Foundation donors. We extend our thanks to the many University of Minnesota resources involved in our project: the University of Minnesota Genomics Center provided services for Sanger and next-generation sequencing; the Flow Cytometry Resource and other services of the Masonic Cancer Center (which is supported by NIH P30 CA77598); the Mass Cytometry Shared Resource at the University of Minnesota which is supported by the Office of the Vice President for Research at the University of Minnesota, and the Minnesota Supercomputing Institute.

Funding Information:
D.A. Largaespada reports grants from American Cancer Society during the conduct of the study; grants from Genentech, Inc., other support from Luminary Therapeutics, Inc., Recombinetics, Inc., Makana Therapeutics, Inc., ImmuSoft, Inc., Styx Biotechnologies, Inc., and NeoClone Biotechnologies International; personal fees from Bio-Techne, Inc., outside the submitted work. No disclosures were reported by the other authors.

Publisher Copyright:
©2022 American Association for Cancer Research.


  • Humans
  • Leukemia, Myeloid, Acute/genetics
  • Oncogenes
  • Oncogene Proteins/genetics
  • Cell Proliferation
  • Cell Line

PubMed: MeSH publication types

  • Journal Article
  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't


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