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Blood, Vol. 92 No. 3 (August 1), 1998:
pp. 1044-1054
Entry and Trafficking of Granzyme B in Target Cells During Granzyme
B-Perforin-Mediated Apoptosis
Michael J. Pinkoski,
Marita Hobman,
Jeffrey A. Heibein,
Kevin Tomaselli,
Feng Li,
Prem Seth,
Christopher J. Froelich, and
R.
Chris Bleackley
From the Department of Biochemistry, University of Alberta, Edmonton,
Alberta, Canada; IDUN Pharmaceuticals, La Jolla, CA; Medicine Branch,
National Cancer Institute, National Institutes of Health,
Bethesda, MD; the Department of Medicine, Evanston Hospital,
Northwestern University, Evanston, IL.
In the widely accepted model of granule-mediated killing by
cytotoxic lymphocytes, granzyme B entry into the target cell is facilitated by the pore forming molecule, perforin. Using indirect immunofluorescence and also direct visualization of fluorescein isothiocyanate (FITC)-conjugated granzyme B, we demonstrate
internalization in the absence of perforin. Induction of the lytic
pathway, however, required a second signal that was provided by
perforin or adenovirus (Ad2). The combination of agents also resulted
in a dramatic relocalization of the granzyme. Microinjection of
granzyme B directly into the cytoplasm of target cells resulted in
apoptosis without the necessity of a second stimulus. This suggested
that the key event is the presence of granzyme B in the cytoplasm, and
that when the enzyme is internalized by a target cell, it trafficks to
an intracellular compartment and accumulates until release is
stimulated by the addition of perforin. We found that the proteinase
passed through rab5-positive vesicles and then accumulated within a
novel compartment. On the basis of these results, we propose a new
model for granzyme-perforin-induced target cell lysis in which
granzyme B is subjected to trafficking events in the target cell that
control and contribute to cell death.
© 1998 by The American Society of Hematology.
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