Urokinase-type plasminogen activator receptor (uPAR) binds
the urokinase-type plasminogen activator (uPA) and
facilitates a proteolytic cascade focused at the cell surface.
More recently, uPAR was recognized as a multifunctional
protein that, through its interactions with integrins, initiates
signaling events that alter cell adhesion, migration and
proliferation. Results obtained recently have led to new
insights into the structural aspects of uPAR interaction with
integrins, provided a more detailed description of the
signaling pathway they induce, and determined that uPAR
signaling plays a role in cell migration and tumorigenicity.
Rochelle Belfer Chemotherapy Foundation, Division of Medical
Oncology, Department of Medicine, Box 1178, Mount Sinai School of
Medicine, New York, New York 11029, USA
Current Opinion in Cell Biology 2000, 12:613–620
0955-0674/00/$ — see front matter
© 2000 Elsevier Science Ltd. All rights reserved.
amino-terminal fragment of uPA
extracellular regulated kinase
focal adhesion kinase
mitogen-activated protein kinase
myosin light chain kinase
p38-MAPK/stress-activated protein kinase 2
urokinase-type plasminogen activator
Results of studies during the past five years have provided
evidence that the urokinase-type plasminogen activator
receptor (uPAR), a glycosyl phosphatidylinositol (GPI)-
linked protein engages in multiple lateral protein–protein
interactions . These results suggested the possibility of a
novel role for uPAR, in addition to its well established func-
tion as a ‘facilitator’ of the cell-surface-based plasminogen
activation that generates a proteolytic cascade important for
matrix degradation in tumor invasion and tissue remodeling
[2,3]. The first hint that uPAR may fulfill multiple roles came
from the studies showing that human myeloid HL60 cells
that overexpress uPAR and uPA are stimulated to adhere to
the substratum and that the adhesion is uPA-dependent .
Another important milestone was the discovery that binding
of uPA, or its amino-terminal fragment (ATF), to uPAR at the
cell surface stimulated cell migration [5,6]. This response
required transmembrane signaling, which could not be
mediated directly through a GPI-linked protein such as
uPAR. There were also reports that uPA treatment can pro-
duce a moderate stimulation of DNA synthesis in some cells
in culture [7,8]. All of these observations supported, albeit in
an indirect way, the notion that by interacting with extracell-
ular domains of transmembrane proteins uPAR is somehow
able to transmit an intracellular signal. A flurry of studies,
completed in the past few years, focused mainly on the
exploration of cytoplasmic proteins that are known to partic-
ipate in signal transduction cascades and thus may form a
complex with uPAR and a putative transmembrane receptor
[9–12]. Using coimmunoprecipitation with anti-uPAR anti-
bodies these studies revealed a puzzling array of associated
proteins, including Src family and other kinases [9,13] but
did not provide any clues as to the nature of the transmem-
brane partners of uPAR that might connect it with the
More recent studies have focused on identifying these
uPAR-associated proteins [10,14–16,17••–19••], as well
as on testing the signaling pathways they evoke
[10,13,17••,18••,20••] and, to a lesser degree, on linking those
changes with biological outcomes such as adhesion, migration,
differentiation and growth [17••–20••,21•,22,23••]. The
results of these studies can be integrated most simply into a
working model that involves uPAR interaction with the β1,
β2 or β3 family of integrins, activation of several groups of
intracellular kinases, and convergence on a specific mito-
gen-activated protein kinase (MAPK) pathway, the
extracellular regulated kinase (ERK) 1/2 pathway. This
scheme does not, however, incorporate all the findings and
it contains many missing links in the signaling cascade and
some uncertainties as to the nature of the uPAR interactions
with other membrane receptors. These uncertainties stem
from the fact that uPAR is anchored through a GPI-linker
and as such may reside in microdomains of the plasma mem-
brane called low density lipid rafts, which contain, in
addition to cholesterol, sphingolipids, glycosphyn-
golipids  and a large collection of proteins . The rafts
are insoluble in non-ionic detergents at low tempera-
ture — the precise conditions often used to lyse cells before
coimmunoprecipitation experiments for detection of uPAR
partners. Proteins that merely coexist in the rafts may some-
times be mistaken for true protein–protein complexes. This
dilemma has not been completely resolved by immunocyto-
chemical colocalization because of its relatively low power of
resolution. In several instances, however, resonance energy
transfer (RET) was observed between uPAR and several
integrins indicating that they are within ~7 nm of each other
and may physically interact . In addition, in more recent
experiments, a peptide homologous to integrin sequences
suspected to be a uPAR-binding site  was identified and
shown to disrupt uPAR–integrin interactions [19••].
Overall, many lines of evidence support the notion that
uPAR is a multifunctional surface receptor with signaling
Urokinase receptor and integrin partnership: coordination of
signaling for cell adhesion, migration and growth
Liliana Ossowski* and Julio A Aguirre-Ghiso†
and adhesive capabilities [1,12,26]. Its expression on the
surface of different cells varies. For example, in contrast to
normal cells, malignant tumor cells usually overexpress
uPAR [6,27–29]. In addition, uPAR’s function as a signaling
molecule can be induced through a conformational change
effected by ligand binding. Alternatively, uPAR, a three-
domain protein, can undergo a limited cleavage which
exposes a sequence between domains 1 and 2 that was
shown to induce signaling and chemotaxis [30–33]. This
plasticity of uPAR and its dynamic interactions with
adapter proteins may be responsible for the observed mul-
tiplicity of signaling pathways and biological outcomes
mediated bu uPAR. The protease-generating function of
uPAR, together with its signaling functions, enable it to
coordinate tumor cell adhesion and invasive migration
through the extracellular matrix (ECM) and to determine
whether tumor cells will, or will not, grow in vivo [18••].
This review focuses on novel aspects of uPAR interactions
with integrins, the signaling pathways that they generate
and the effect of these interactions on the biology of both
normal and tumor cells.
Structural properties of uPAR required for its
uPAR, which was discovered and cloned 15 years ago ,
is synthesized as a 313 residue polypeptide that is folded,
through disulfide bonding, into three homologous
repeats, known as domains 1, 2 and 3 (; Figure 1).
Interdomain connecting peptides, and especially the
peptide that connects domains 1 and 2, were shown to
have chemotactic properties . uPAR contains five
potential glycosylation sites and it is known to be heavily
glycosylated . Its carboxy-terminal portion (in domain 3)
is processed to add a GPI-anchor, which targets the
receptor to the outer leaflet of the plasma membrane
lipid bilayer. The amino-terminal portion of the protein
(domain 1) provides the uPA-binding site, but there is
evidence that residues in domain 3 also participate in the
assembly of the ligand-binding site and that domains 2
and 3 increase the affinity of uPA binding to domain 1
[35,37•,38]. On the surface of cells, uPAR exists in either
the three-domain form, which is capable of binding uPA,
or the two-domain form (devoid of domain 1), which does
not bind uPA [30,31].
Cell-to-cell contact and extracellular matrix
Migration in culture
•Proliferation in culture
•In vivo tumorigenesis
Migration in culture
Current Opinion in Cell Biology
Alternative models for uPA–uPAR-initiated signal transduction and its
biological consequences. (a) In tumor cells, overexpressed uPA bound
to uPAR interacts with α5β1 integrins (α,β) with high frequency, keeping
a large proportion of the integrins in an active state. Clustered active
uPA–uPAR–α5β1 complexes bind FN and initiate both intracellular
signaling and extracellular FN fibril formation. Through a caveolin-
independent mechanism the active uPA–uPAR–α5β1–FN fibril complex
triggers a powerful activation of the Mek-ERK pathway. The presence of
FN fibrils, along with cytoskeleton reorganization through an unknown
pathway, keeps the growth suppressive pathway p38MAPKinhibited.
Under these conditions the balance is in favor of the ERK pathway and
when ERK activity reaches a threshold level, tumor cells begin to
proliferate in vivo (in animal models). (b) In kidney epithelial or vascular
smooth muscle cells, ligand-clustered integrins and caveolin
oligomerization release Src family kinases, allowing them to be activated.
uPAR, complexed with caveolin and signaling molecules, enters the
cluster, binds to a small number of integrins, bringing the signaling
molecules into the cluster. FAK and other signals (probably a
phosphatase(s) and Fyn) are activated. This leads to ERK activation,
which, combined with active FAK and cytoskeleton changes, is required
for matrix-dependent migration. This (caveolin-dependent) mechanism
may be also involved in uPAR-Mac1-dependent migration of leukocytes
in vivo. (c) In mammary tumor cells, uPA binding to uPAR activates,
through a still unknown partner molecule, FAK and Src phosphorylation,
which leads to activation of the classic mitogenic pathway Shc-Ras-Mek-
ERK. ERK, directly or indirectly, activates MLCK, which phosphorylates
myosin regulatory light chain (not depicted) and initiates cytoskeleton
contraction and cell migration in an integrin- and matrix-dependent
fashion. According to the levels of uPAR expression in the different
studies it is possible that (b) and (c) models require a small or ‘normal’
number of uPAR molecules, whereas model (a) requires a high number
of surface uPARs, which is observed in highly malignant tumor cells.
Generation of signals through uPAR
Effect of ligand binding and limited proteolysis on
It has been assumed that to initiate signaling, uPAR under-
goes a conformation change. This conclusion was based on
the observation that binding of uPA to uPAR induced
adhesion, migration, proliferation, and maintenance of
differentiation programs preceded by a signaling event and
activation of genes. Only a few studies have directly
addressed the question of whether conformational change
is required for signaling. One approach compared the func-
tion of a mutant uPA, in which the amino-terminal 24 amino
acids were substituted with a random stretch of amino
acids, to that of a wild-type uPA . Both forms of uPA
showed similar binding to uPAR, but only the wild-type
uPA induced cell migration. As uPA must induce intracellu-
lar signalling to induce migration, it can be inferred that
only when a conformational change that exposes a region
responsible for interaction with an adapter protein is affected
can uPAR be effective in signaling. .
It has been shown that chemotaxis of monocytic THP-1
cells can be induced not only by uPA or its ATF, but also
by the two-domain form of uPAR, generated by proleolytic
cleavage of domain 1 [30,31]. In these experiments soluble
uPAR (suPAR) was not active. . Cleaved uPAR (domains 2
and 3), as well as uPA or ATF, induced activation of a
Src-like tyrosine kinase Hck and pertussis toxin sensitive,
heterotrimeric G proteins [30,31] (Table 1). Further stud-
ies revealed that cleaving domain 1 exposed a linker region
that connects domains 1 and 2 in intact uPAR. This pep-
tide contains the sequence SRSRY. A synthetic peptide
that contains this sequence can stimulate chemotaxis of
THP-1 cells, uPAR-deficient macrophages and NIH3T3
cells at 0.1 pM concentration , suggesting that this may
be the cis-interacting epitope of uPAR
In MCF-7 breast carcinoma cells, not only the connecting
peptide but the intact suPAR-induced cell migration .
The effect on migration of uPA was greater than that of
suPAR or the peptide. Similarly, we observed that in an
epithelial cancer cell line, the suPAR-induced ERK acti-
vation was less than that induced by uPA [18••]. The fact
that intact suPAR, in which presumably the connecting
peptide is buried, retains activity may mean that the con-
necting peptide is not the main epitope responsible for
interaction with integrins. Alternatively, since the puri-
fied, connecting peptide can stimulate chemotaxis,
without the participation of the rest of uPAR-molecule, it
is likely that it may interact with another receptor, yet ini-
tiate a similar pathway of Hck activation. The latter
possibility was supported by the recent observations that
uPAR lacking domain 1 (and thus presumably with an
accessible connecting peptide) does not coimmunoprecip-
itate with β1 integrin , weakening the argument for
the peptide as the main interacting epitope (see Update).
Interactions of uPAR with adapter proteins
The current hypothesis is that a change in uPAR confor-
mation allows it to interact with a signal-transducing
adapter protein. As stated earlier, integrins of the β1, β2
and β3 families have been shown to functionally interact
with uPAR upon binding to specific ECM ligands .
These interactions are dynamic, since, as shown by the
RET technique in neutrophils, uPAR colocalizes with CR3
integrin in stationary cells but it segregates away from this
Urokinase receptor and integrins Ossowski and Aguirre-Ghiso 615
Signaling modules activated by uPA–uPAR or the uPA–uPAR–integrin complex and the biological outcomes.
Signaling modules Cell types studiedBiochemical and biological outcomesReferences
FAK-Cas-PaxilinLNCaP, ECMigration [48,49]
FAK, Src, ERKMCF-7, HKC293 Adhesion, migration [17••,40]
Ras-ERKMCF-7, MCF-7-uPARMigration [20••]
Mek-ERKMCF-7, MCF-7-uPAR, HEp3, HKC293Adhesion, migration, tumor growth in vivo
Hck-Fgr U937Chemotaxis, adhesion, differentiation[31,45]
THP-1, VSMCChemotaxis, differentiation, cytoskeleton dynamics [31,44]
HT-1080c-fos, PAI-2 expression
P38HT-1080, HEp3 c-fos, PAI-2 expression, tumor growth in vivo
Jak/Stat familyVSMC, ECTranscription, DNA binding, distribution in cell extensions[10,11]
Cells: EC, endothelial cells; HEp3, human squamous carcinoma;
HKC293, human kidney embryonic; HT-1080, human fibrosarcoma;
LNCaP, human prostate cancer; MCF-7 human mammary carecinoma;
MCF-7-uPAR, MCF-7 overexpressing uPAR; PC12,
phaeochromocytoma; THP-1, human monocytic; U937, human
myelo-monocytic; VSMC, vascular smooth muscle cells.
receptor into the advancing edge lamellipodia during
migration, leaving CR3 in the uropod .
The simultaneous presence of both uPAR and integrins on
the cell surface prompts the question of what promotes
and what disrupts their interactions. Two somewhat oppos-
ing scenarios have been proposed (Figure 1b). In one,
integrins are thought to be activated in a caveolin-depen-
dent fashion by ligand binding before they interact with
uPAR [17••,18••]. The second scenario proposes that these
interactions may be caveolin independent. A series of stud-
ies have identified a specific uPAR-binding site on
αM subunit of the CD11b/CD18 integrin. It has been pro-
posed that the amino-terminal portion of the αM integrin
subunit folds in a β-propeller-like structure formed by
repeating units (W1–W7) of anti-parallel β-sheets, inter-
rupted by I-domain inserted between W2 and W3. The sites
for uPAR binding have been mapped to the W4 ‘blade’ of
the β-propeller of αM integrin not far from the I-domain,
which is the ligand (fibrinogen)-binding domain [14,19••].
This proximity may explain the inhibition by suPAR of fib-
rinogen binding [19••]. In contrast, ample evidence
indicates that uPAR promotes normal functions of β1 and β2
integrins [16,18••,22], posing an apparent paradox to the
finding that it inhibits ligand binding. An explanation for
this paradox may be found in the recent report [17••] that
proposes that integrin clusters contain only a few uPAR mol-
ecules and that by binding to the integrin in a ligand-like
fashion, uPAR enriches the cluster with signaling molecules
at the same time as preventing the few uPAR-bound inte-
grins from binding to their natural ligand [17••] (Figure 1b).
What it means, in effect, is that some capacity for ligand
binding is sacrificed for the enrichment of the integrin clus-
ters with signaling molecules (Figure 1b). It also appears,
that, in some cells, an important determinant of uPAR–inte-
grin function is caveolin, a protein that is known to
oligomerize, associate with Src family kinases [41,42], and
coimmunoprecipitate with β1 integrins and uPAR
[14,17••,43]. Several experiments point to the possibility that
the level of caveolin influences the outcome of uPAR–inte-
grin interactions. For example, human kidney cells 293
adhere well to fibronectin but this adhesion is abolished by
expression of uPAR . Overexpression of caveolin in these
cells completely restored adhesion to fibronectin [17••]. Wei
et al. [17••] propose that activation of integrins in response to
ligand binding results in oligomerization of caveolin. This
releases Src kinases, which become activated, while at the
same time uPAR moves into the integrin–caveolin–Src
kinase complexes ‘enriching’ them with caveolin and its
associated signaling molecules and stimulating integrin func-
tion [17••]. A peptide identical to the uPAR-binding
sequence on the W4 blade of αM integrin disrupts
uPAR–integrin interaction and inhibits cell migration [19••],
providing additional support for the notion that binding of
uPAR provides a positive stimulus for integrin function.
The second scenario indicates that the need for caveolin is
not universal, as demonstrated by the results obtained with
the tumorigenic human carcinoma cells, HEp3. These
cells express very high level of uPAR, adhere very strongly
to fibronectin through the α5β1 integrin but do not express
caveolin [18••] (Figure 1a). Adhesion of these cells to
fibronectin induces ERK activation. As proposed by the
first scenario, in high uPAR expressing cells such as HEp3,
because of high frequency of uPAR–α5β1 integrin interac-
tions, the integrin should be inactive. The opposite has be
found to be true; α5β1 are active until uPAR is reduced to
approximately 20% of that present in parental HEp3 cells.
Simultaneously, the fibronectin-induced signal to ERK is
drastically reduced [18••]. Therefore, in this model system,
it is the abundance or dearth of uPAR that determines the
state of integrin activation simply by increasing the chance
of interactions between the two proteins (Figure 1a).
Whether another scaffolding protein is substituting for
caveolin in cells, or whether mechanisms of integrin
activation by uPAR are cell specific, is currently unknown.
Functional consequences of activation of
intracellular pathways by the
As discussed earlier, the understanding of the details of
uPA–uPAR–integrin interactions and the precise way they
assemble to generate signals are still missing. . In contrast,
there is substantial evidence that these interactions acti-
vate signaling pathways and adhesion or migration events.
Studies that have focused on mapping the specific path-
ways and testing their biological consequences are more
relevant than a mere identification of signaling proteins
associated with the uPAR molecule. (The summary of
uPAR-activated pathways is shown in Table
Coimmunoprecipitation of detergent-lysed cell proteins
with anti-uPAR antibodies has identified a multitude of
potential candidates for signaling [9–11], among them are
a number of members of the Src kinase family [9,13]. In
THP-1 cells treated with uPA, ATF or uPAR-devoid of
domain 1, a Src family kinase Hck and a trimeric pertussis
toxin sensitive G protein were activated, leading to their
enhanced migration [30,31]. A similar activation pathway
seems to operate in smooth muscle cells and to induce
reorganization of stress fibers into cortical F-actin, relocat-
ing c-Src, αvβ3- and β1-integrins to the leading edges of
cells migrating on vitronectin .
Another study  examined whether uPA-binding to
uPAR changed the composition of the uPAR complex or
changed the state of activation of uPAR-associated kinases.
These authors have shown that Src kinase Hck is activated
within minutes of treatment with high concentration
(50 nM) of uPA, and there is also a simultaneous activation
of ERK, p38 and then induction of c-fos mRNA level and
activation of PAI-2. However, neither a transducing partner
has been identified nor a firm connection been established
between the activation of Hck and activation of ERK.
Signaling through uPAR also plays a role in myeloid cell
differentiation. U937 cells ‘primed’ for differentiation with
TGFβ or VitD3 adhere to the substratum and progress
Cell-to-cell contact and extracellular matrix
further along the differentiation pathway when treated
with uPA or ATF, a process that leads to inhibition of Hck
and Fgr tyrosine kinases . In contrast, ‘non-primed’
cells, which do not differentiate, are stimulated to migrate,
and not to adhere, upon treatment with uPA or ATF, and in
these cells Hck is activated . Thus, the context in
which the uPA signal is received regulates the switch
between active and inactive states of Hck and Fgr kinases
and establishes whether myeloid cells will differentiate
further or remain undifferentiated and more motile.
As integrins have been identified as the transmembrane
partners of uPA/uPAR, it was expected that the same sig-
naling pathways linked to their activation by other means
might be functional in uPA/uPAR activation. Indeed, it
appears that the signaling converges on activation of the
MAPK-ERK pathway, although there are variations and
missing details [17••,18••,20••,40]. For example, MCF-7
cells with very low number of uPAR receptors
(~3000 sites per cell) are stimulated to migrate at low nM
concentrations of uPA or ATF fragment. This is associated
with a very transient (1 min) stimulation of MAPKs
ERK1/2 . It is known that the Mek-ERK pathway
can, through phosphorylation of myosin light chain
kinase (MLCK), enhance migration . In MCF-7 cells
overexpressing uPAR, uPA-stimulated migration was also
linked to ERK activation and MLCK phosphorylation
[20••]. The effect was specific for vitronectin and its
receptor ανβ5 integrin [20••], although the involvement
of the uPAR–integrin interaction in this system remains
unknown. In HEp3 cells, uPA binding to uPAR induces a
strong, persistent, fibronectin-dependent activation of
Mek and ERK, which is crucial for the growth of these
cells in vivo [18••]. A more recent report  has shown
that uPA-induced Ras-ERK migration is regulated
upstream by activation of the focal adhesion kinase
(FAK), followed by the classic assembly of kinase and
docking proteins. The Shc–SOS complex was very tran-
sient (1–2.5 min), which matched the previously shown
transient activation of ERK . It is intriguing that such
a transient signal can influence such a long lasting event
as cell migration, suggesting that a parallel and more sus-
tained pathway activated by uPA/uPAR may be activated.
Transient activation of FAK and phosphorylation of Cas
by uPA treatment have been shown in LNCaP cells trans-
fected with uPAR, although the implications of this
pathway for migration in LNCaP cells has not been
established [48,49]. In endothelial cells treated with
50 nM uPA (which is 50 fold greater than the uPA Kd)
induction of FAK, paxillin, p130cas, and ERK pathway
has been shown, but the mechanism through which uPAR
activates FAK is not known .
Several other signals were identified as a result of uPA
binding to uPAR. In endothelial and smooth muscle cells,
uPA induced activation of the Jak/Stat pathway [10,11], but
these studies have not yet progressed beyond the identifi-
cation of the signaling molecules. Koshelnick et al.  and
Dumler et al.  found that uPA-induced active Jak1
and Tyk2 codistribute with uPAR when the cells are
migrating into a wound, whereas the Src kinase, although
present, is randomly distributed . In another study,
activation by uPA induced activation of Stat1, 2 and 4 but
neither biological effects nor the transmembrane
adaptors were examined [50•].
Perturbing the uPAR receptor either by uPA binding or by
aggregation through specific crosslinking antibodies evokes
several additional signaling events. Two reports [51,52]
tested the effect of uPA or crosslinking antibodies on the
level of intracellular Ca2+in U937 cells. In both cases, Ca2+
release from the intracellular stores was documented. It is
of interest that in this monocytic cell line, the uPAR-depen-
dent Ca2+increase appeared to be independent of
uPAR-association with the β2 integrin. This is in contrast to
the role of CR3 integrin in Ca2+release in neutrophils .
These examples indicate that even in cells of hematopoietic
origin (neutrophils and monocytes) the final outcome can
be achieved through different mechanisms.
The most likely pathway for signaling triggered by
uPA–uPAR–integrins deduced from the currently available
evidence is the Mek-ERK pathway with upstream feeding
signals derived from Src, Src-like tyrosine kinases (Hck
and Fgr) and FAK. The cell types where these pathways
are activated may have different biological outcomes, such
as differentiation in myelomonocytic cells, motility in
endothelial and vascular smooth muscle cells or motility
and growth in epithelial or mesenchymal tumor cells.
Role of uPA/uPAR signaling in cell migration
and growth in vivo
uPAR-dependent migration of leukocytes in vivo
The role of the signaling properties of the uPA–uPAR com-
plex in vivo remains relatively unexplored, although the
existing data imply a crucial role for this function of uPAR.
Activated leukocytes express uPA and uPAR, which
enhance their adhesion in culture . To study the role of
the uPA–uPAR complex in vivo, leukocyte recruitment to
sites of inflammation in the peritoneum was studied .
In uPAR null mice, leukocyte recruitment was reduced by
80%, and the residual migration was only marginally affected
by the administration of antibodies against ICAM-1 and
LFA-1. In contrast, in wild-type mice, the recruitment of
leukocytes was effectively blocked by these antibodies,
down to the level of cells that migrated in uPAR null mice.
The importance of intact uPAR in this process is under-
scored by an experiment in which adhesion of leukocytes to
endothelium in culture was examined. When uPAR was
cleaved from the surface of cells with PI-PLC (phos-
phatidyl inositol phospholipase C), suPAR, but not the
two-domain uPAR, restored their adhesion  (see
Update). This supports the conclusion that an intact uPAR
molecule, probably by activating β2 integrin, is required for
in vivo transendothelial migration.
Urokinase receptor and integrins Ossowski and Aguirre-Ghiso 617
To test the role of uPAR occupancy in cellular migration,
in vivo influx of neutrophils to the lung was induced by a
chemoatractant (KC, a murine homologue of IL-8) in
wild-type, uPA null and uPAR null mice . KC stimu-
lated neutrophil influx into the lungs in all three types of
mice. When mice were pre-treated with a proteolytically
inactive ligand (a fusion protein of the growth factor
domain of uPA (amino acids 1–48, which compete with
endogenous uPA binding and the human Fc domain of
IgG (GFD-Fc)) of uPAR , the influx of neutrophils
was strongly impaired in wild-type and uPA null mice but
not in uPAR null mice. These results suggest that the
complete absence of uPAR does not influence neutrophil
migration, but when present, uPAR bound to an inappro-
priate ligand (GDF instead of uPA) may lose its normal
function. Moreover, uPAR occupancy by a ligand that is
cleared inefficiently from the cell surface may alter
uPAR-dependent integrin function ( and Update).
Signaling through uPAR in cancer: a switch between
tumor growth and dormancy
A striking feature of malignant solid tumors is uPAR over-
expression [2,29]. Although the role of uPAR in zymogen
activation is well understood, the signaling role of the high
number of receptors in tumor cells is almost unknown. It
was discovered that by downregulating uPAR expression
about fourfold HEp3 carcinoma cells enter a state of dor-
mancy in vivo [18••,28]. The analysis of the mechanism
revealed that the overexpressed uPA–uPAR complex
interacted with α5β1 integrin at a high frequency, leading
to a powerful and persistent activation of the Mek-ERK
pathway upon binding to fibronectin (FN). When uPAR
was downregulated or the uPAR–α5β1 interaction was dis-
rupted, the ERK pathway became deactivated and the
cells arrested in G0/G1phase of the cell cycle in vivo and
tumor dormancy was triggered [18••].
Cells with high uPAR levels, which resulted in active
α5β1, proficiently assembled FN fibrils (JA Aguirre-Ghiso
et al., unpublished data). Downregulation of uPAR and
subsequent inactivation of α5β1 in dormant cells caused a
total loss of the ability to assemble FN fibrils. More impor-
tantly, we found that the function of uPAR, which was
required to maintain the fibrils, was crucial for keeping the
p38MAPKpathway (proapoptotic/growth suppressive) in an
inactive state. Thus, uPAR overexpression uses integrins to
generate a mitogenic signal by regulating two opposing path-
ways: activation of ERK and inhibition of p38MAPK[18••]
(JA Aguirre-Ghiso et al., unpublished data).
A confirmation of the growth promoting role of uPAR
was recently obtained in a clinical study [55,56]. Patients
with gastric cancer were subjected to serial bone marrow
biopsies at the time of surgery for the removal of the
tumor and every six months thereafter. Their bone mar-
row cells were stained for cytokeratins and uPAR. In
patients in which the original sample contained uPAR
positive cells, the number of such cells increased in
subsequent samplings and the patient had a shorter
relapse-free interval. In contrast, when the original bone
marrow sample had few or no uPAR positive cells, the
number of tumor cells (cytokeratin positive) in subse-
quent samples was either stable or reduced, and the
patients had a longer relapse-free period. These results
suggest that the presence of uPAR on cancer cells in the
bone marrow may induce a growth-promoting signal,
ending the dormancy period of the disease.
The unexpected idea that the urokinase receptor, a
GPI-linked protein, can serve the function of a signaling
protein is becoming better established. Currently, its best
studied partners are members of the integrin family, but
other partners can not be excluded. It is possible that a rel-
atively ‘mobile’ membrane protein like uPAR can, by
multiple interactions, increase the cell repertoire of regula-
tory mechanisms. Current evidence supports the notion
that the classical MAPK signaling pathway of integrins is
also the major uPAR-activated pathway. The possibility of
uPAR influencing multiple biological events, such as
migration, adhesion, tumorigenicity and differentiation, by
lateral interactions with extracellular domains of mem-
brane proteins is novel and exciting. As uPAR appears to
function as a regulator of integrin activity, its level of
expression on the surface of cells, its molecular integrity,
and level of glycosylation may have a profound regulatory
effect. Much, however, remains to be determined. Is ERK
always in the center of the signaling cascades? How can
activation of Jak/Stat be integrated into this working
model? Are there scaffolding proteins in addition to caveo-
lin, or can uPAR function through a different mechanism?
Are there potential therapeutic implications for cancer,
inflammatory and vascular disease? The recent rapid rate
of discoveries in this field will undoubtedly lead to answers
to many of those questions.
Recent reports provide support for the role of uPAR in reg-
ulating integrin function during leukocyte adhesion  and
recruitment to injury sites in response to bacterial infection
. May et al.  reported that engagement of VLA-4
(α4β1 integrin) in HL60 or monocytic leukemia U937 cells
has a dominant-positive function by activating β2-integrin-
dependent adhesion to endothelial cells. Most importantly,
dominant function of VLA-4 is mediated by uPAR, as its
removal from the cell surface dramatically reduces VLA-4
activation of β2 integrins. In agreement with the hypothesis
that full length uPAR (but not uPAR devoid of domain 1)
contains the epitope(s) required for integrin interaction,
only full length soluble uPAR restored integrin function
after PI-PLC treatment, whereas treatment with
D2+D3-suPAR did not restore integrin function .
Recruitment of neutrophils to sites of inflammation upon
bacterial infection is crucial for a proper immune response.
Gyetko et al.  recently showed that while recruitment
Cell-to-cell contact and extracellular matrix
of neutrophils to the lung followed infection with
Pseudomonas aeruginosa was rapid in uPAR+/+mice, it was
drastically reduced impaired in uPAR–/–mice for up to four
to eight hours. In uPAR+/+mice this response was inhibited
by pre-inoculation with anti-CD11b antibodies. In addi-
tion, while clearance of the bacteria from the lungs was
highly efficient in uPAR+/+mice, uPAR–/–were unable to
clear P. aeruginosa from the lungs as evidenced by a higher
bacterial content (CFU) in bronchioalveolar lavage .
These results support the crucial role for uPAR in
transendothelial migration in vivo, which depends also on
β2 integrins [22,23••], towards the tissue parachyma to
proceed with bacterial clearance during infeciton.
This work was supported by a US Public Health Service Research Grant
(CA-40758) and the Samuel Waxman Cancer Research Foundation. This
article was not meant to be a comprehensive review of the existing literature
and because of space constraints not all the relevant work has been
included. We apologize in advance for what were not intentional omissions.
Some topics, such as the role of uPAR in adhesion to vitronectin, have not
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Cell-to-cell contact and extracellular matrix