Serial sections were either hybridized with a e35S-UTP antisens riboprobe for the detection of EMAP II transcripts (dark field A-C; bright field D-F) or stained for DNA fragmentation (G-I). Shown are sections of the yolk sac (A, D, G), the eye (B, E, H), and the trigeminal ganglion (C, F, I). Full size image available through contact |
Endothelial monocyte-activating polypeptide II (EMAP II) was
originally purified from supernatants of Meth A fibrosarcoma
cells due to its ability to induce tissue factor, the
initiator of coagulation, on endothelial cells. EMAP II has
been characterised as a proinflammatory cytokine capable of
stimulating the chemotactic migration of monocytes and
neutrophils as well as of activating endothelial cells and
monocytes to become macrophages. In vivo, injection of EMAP
II into the mouse footpad results in an inflammatory swelling
response characterised by a cellular infiltrate and edema.
Cloning of the EMAP II-cDNA revealed that EMAP II is
translated as a precursor protein which is processed to
become the biologically active mature form that is
subsequently released from the cell. Since both the human and
the mouse EMAP II precursor proteins are cleaved after an
aspartic acid residue, the EMAP II cleavage site resembles
target sites of caspases, a family of proteases with a
pivotal role in apoptosis, also known as programmed cell
death.
Little was known about the expression of EMAP II or the
regulation of its cleavage. Scientists in W. Risau’s
department at the Max Planck Institute for Clinical and
Physiological Research in Bad Nauheim have analyzed the
expression pattern of the EMAP II gene during embryogenesis
of the mouse and found that the EMAP II messenger-RNA (mRNA)
is predominantly expressed at sites of tissue remodelling.
Since it is known that programmed cell death is a necessary
requirement for tissue remodelling, mouse embryos were
analyzed for the presence of apoptotic cells. A high rate of
apoptosis was detected in tissues with a strong EMAP II mRNA
expression. The removal of dead cells during apoptosis
requires macrophages and an accumulation of macrophages in
areas of EMAP II expression and apoptosis could be observed.
As the expression pattern of EMAP II during mouse development
suggested a role for programmed cell death in the regulation
of EMAP II, the effect of apoptosis on the posttranslational
processing of the EMAP II precursor protein was investigated in
vitro. It could be shown that following the induction of
apoptosis, the EMAP II precursor protein was cleaved and the
mature EMAP II released from the cell, whereas the induction
of necrotic cell death had no effect on the processing of the
EMAP II precursor. The release of mature EMAP II following
the induction of apoptosis could be abrogated by a specific
tetrapeptide inhibitor which competes with the cleavage site
of the EMAP II precursor. Further experiments using other
peptide inhibitors indicated that EMAP II is cleaved by a
caspase-like enzyme.
This study demonstrates that, during embryonic development,
the EMAP II mRNA colocalizes with sites of programmed cell
death and that the posttranslational processing of the EMAP
II precursor is induced by apoptosis. Since the mature EMAP
II is a chemoattractant for macrophages, its release
following apoptosis suggests that EMAP II recruits
macrophages to sites of apoptosis. So far, no other mechanism
has been described which can explain the accumulaion of
macrophages at sites of cell death during embryonic
development. Taken together, these results imply that the
coordinate program of cell death includes activation of a
capase-like activity that initiates the processing of a
cytokine responsible for macrophage attraction to the sites
of apoptosis.
Original paper:
Knies, U. E., Behrensdorf, H. A., Mitchell, C. A.,
Deutsch, U., Risau, W., Drexler, H. C. A. and Clauss, M.:
Regulation of endothelial monocyte-activating polypeptide II
release by apoptosis. PNAS 95 (21):
12322 - 12327 (1998).
Journal
Proceedings of the National Academy of Sciences