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Dissection of the c-Kit signaling pathway in mouse primordial germ cells by retroviral-mediated gene transfer

  1. Peter J. Donovan*
  1. *Kimmel Cancer Center, Thomas Jefferson University, Bluemle Life Sciences Building, Room 706, 233 South 10th Street, Philadelphia, PA 19107; †Division of Immunology and Hematopoiesis Oncology Center, Room 1-109, Johns Hopkins University School of Medicine, 1650 Orleans Street, Baltimore, MD 21231; ‡Departments of Neurosurgery, Neurology, and Cell Biology, Memorial Sloan Kettering Cancer Center, Rockefeller Research Laboratories, Room 917B, 1275 York Avenue, New York, NY 10021; and §Molecular Medicine Program, Mayo Clinic and Mayo Foundation, Rochester, MN 55905

  1. Communicated by John W. Littlefield, Johns Hopkins University School of Medicine, Baltimore, MD (received for review November 7, 2001)

Abstract

Establishment of the mammalian germ line is a prerequisite for fertility of the adult animal but we know surprisingly little about the molecular mechanisms regulating germ-line development in mammals. Signaling from the c-Kit receptor tyrosine kinase is essential for primordial germ cell (PGC) growth both in vivo and in vitro. Many downstream effectors of the c-Kit signaling pathway have been identified in other cell types but how these molecules control PGC survival and proliferation are unknown. Determination of the c-Kit effectors acting in PGCs has been hampered by the lack of effective methods to easily manipulate gene expression in these cells. We overcame this problem by testing the efficacy of retroviral-mediated gene transfer for manipulating gene expression in mammalian germ cells. We found that PGCs can be successfully infected with a variety of types of retroviruses. We used this method to demonstrate an important role for the AKT kinase in regulating PGC growth. Such technology for manipulating gene expression in PGCs will allow many of the molecular mechanisms regulating germ cell growth, behavior, and differentiation to be comprehensively analyzed.

Footnotes

    • ↵ To whom reprint requests should be addressed. E-mail: pdonovan{at}springfield.jci.tju.edu.

  • Abbreviations

    • PGC, primordial germ cell

    • KL, Kit ligand

    • ALV, avian leukosis virus

    • MLV, murine leukemia virus

    • MSCV, murine stem cell virus

    • GFP, green fluorescent protein

    • Sl, Steel

    • PI3K, phosphatidylinositol 3-kinase

    • MAPK, mitogen-activated protein kinase

    • MEK, mitogen-activated protein kinase kinase

    • FRAP, FKBP-12-rapamycin-associated protein

    • HA, hemagglutinin

    • Received November 7, 2001.
    • Accepted April 26, 2002.

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