INSIGHTS | PERSPECTIVES
GRAPHIC: KELLIE HOLOSKI/SCIENCEscience.org SCIENCEregulate vascular invasion and initiate BBB
formation ( 2 ). In the adult brain, astrocyte,
neuron, and oligodendrocyte progenitor cell–
derived Wnt7 signaling maintains the BBB
through canonical, Frizzled (Fz) receptor–
and b-catenin–dependent, signaling ( 8 ). In
addition, vascular endothelial signaling uses
a distinct Wnt7 co-receptor complex compris-
ing G protein–coupled receptor 124 (GPR124)
and the glycosylphosphatidylinositol (GPI)–
anchored surface protein RECK (reversion-
inducing cysteine-rich protein with Kazal
motifs) ( 9 – 10 ). Localized RECK-GPR124-
Wnt7 complex activity in brain endothelial
cells stimulates Wnt7 signaling through Fz
receptors.
Crystallographic and biochemical evi-
dence has identified two regions in Wnt
ligands, one toward the amino terminus
(site 1), the other toward the carboxyl ter-
minus and associated also with selective
Fz interactions (site 2) (see the figure) ( 11 ).
A distinct linker domain on Wnt7 ligands
is thought to mediate Wnt7 association
with the RECK-GPR124 complex, binding
to RECK ( 12 ). Martin et al. showed that
amino-terminal Wnt7a, with the linker do-
main, was sufficient to activate Fz signaling
but only when RECK-GPR124 were present,
suggesting a strategy for restricted Wnt
pathway activation.
Screening identified 25-point muta-
tions in Wnt7a that confer similar RECK-
GPR124 selectivity. One of these, K190A
(Lys^190 Ala) was singled out for further
study. Wnt7a-K190A is a sensitive agonist of
RECK-GPR124-Fz5/8 signaling with limited
off-target activity when only the Fz recep-
tors are present. This selective outcome
is critical for therapeutic utility, given the
potential for wide-ranging pathology from
activation of broadly distributed Fz recep-
tors. Although the molecular explanation
for selectivity is unclear, the authors sug-
gest that decreased stability of the mutant
form may affect interactions with Fz recep-
tor complexes.
To address the in vivo actions of Wnt7a-
K190A, the authors used gene editing in
zebrafish to demonstrate that Wnt7aa activ-
ity is required for early vascular ingrowth
and activation of the glucose transporter,
Glut1, indicating initiation of BBB forma-tion. Introduction of Wnt7a-K190A rescued
Wnt7aa mutant phenotypes in a transgenic
model. Comparison of the sites of Wnt7a-
K190A production in relation to the ob-
served rescue suggests long-range activity,
which is potentially at odds with the re-
duced stability and short-range signaling
interpretation from in vitro cell culture
studies. Further analysis in a zebrafish
stroke model showed reduced bleeding and
leakage in wild-type zebrafish ectopically
expressing Wnt7a-K190A, which is consis-
tent with improved BBB activity.
Moving to mouse models of BBB break-
down—brain tumors and stroke—the au-
thors delivered Wnt7a-K190A through ad-
eno-associated virus (AAV) injection into
the circulation, demonstrating broad in-
fectivity of endothelial and nonendothelial
associated cell types in the brain. Wnt7a-
K190A showed no off-target (nonvascularendothelial) activation of Wnt signaling
in the brain. Implantation of glioblastoma
tumor cells resulted in a breakdown of
BBB properties and vascular leakage. The
accompanying reduction in Wnt7-target
gene expression in vascular endothelium
was ameliorated by infection with AAV
encoding Wnt7a-K190A. Restoration of
Wnt-signaling and BBB properties reduced
tumor size, whereas inhibiting canoni-
cal Wnt signaling, with the secreted Wnt-
inhibitor Dickkopf-1 (DKK1), enhanced
tumor growth. Wnt7a-K190A also showed
beneficial effects in a cerebral artery occlu-
sion model of stroke. The infarct volume
was reduced, and post-stroke reperfusion
injury, which is linked to vascular leakage,
was limited, pointing to improved BBB
function, which is consistent with studies
transiently stabilizing b-catenin ( 9 ).
Whether these beneficial effects reflect a
direct action of Wnt7a-K190A on the BBB
was not addressed. The mechanism is im-
portant given previous reports suggesting
that Wnt responses in the BBB were not al-
tered by stroke ( 13 ). It will be interesting to
determine whether modified Wnt ligands
could be effective for other indications in
which BBB dysfunction has a causal or
early role in pathogenesis. For example,
biomarkers of BBB breakdown showed
better correlation with the emergence of
cognitive impairment in Alzheimer’s dis-
ease patients than did amyloid-b or tau pa-
thology ( 14 ). Overall, Martin et al. make a
strong case for modified Wnt ligands as po-
tential pharmacological agents that could
stimulate specific Wnt receptor–mediated
cellular outcomes in selected cell types. jREFERE NCES AND NOTES- M. D. Sweeney, Z. Zhao, A. Montagne, A. R. Nelson, B. V.
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ACKNOWLEDGMENTS
A.P.M. is supported by grants from the National
Institute of Diabetes and Digestive and Kidney Diseases
(UC2DK126024, DK054364, DK121409, and DK126925).
A.P.M. declares no conflicts of interest. J.K.I. is a cofounder
of AcuraStem and Modulo Bio and serves on the scientific
advisory board of Spinogenix.
10.1126/science.abn7921Wnt7a Wnt7a-K190ASite 1 Site 2
Frizzled
(no GPR124-RECK)Frizzled
(no GPR124-RECK)
ActivationWnt signaling
in multiple cells
and tissuesActivationReduced
e ects
outside CNSGPR124
RECKFrizzledBBBImproved BBB
function after strokeReduced glioblastoma
tumor growthNo activation“Restoration of Wnt-signaling
and [blood-brain barrier]
properties reduced tumor size...”
Engineering Wnt7a for blood-brain barrier–specific activity
A K190A (Lys^190 Ala) point mutation in Wnt7a restricts Wnt7a signaling to cells with the co-receptor complex
GPR124 (G protein–coupled receptor 124) and RECK (reversion-inducing cysteine-rich protein with Kazal
motifs), activating Wnt signaling specifically in endothelial cells of the central nervous system (CNS). When
introduced into mouse brains, Wnt7a-K190A regulates blood-brain barrier (BBB) functions and limits
the pathogenesis of brain tumors and stroke.
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