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Page 10 of 23 Padarti et al. Vessel Plus 2018;2:21 I http://dx.doi.org/10.20517/2574-1209.2018.34

may be critical for the maintenance of cellular architecture as well as regulation of β1-integrin-mediated
signaling . The follow-up studies found that ICAP1α deficiencies result in many osteoblastic defects which
[25]
are the direct results of β1-integrin activation. CCM1 acts as a competitive inhibitor for the interaction of
ICAP1α and β1 integrin. The lack of inhibition of ICAP1α leads to excessive inhibition of β1-integrin which
is thought to cause the leaky vasculature found in CCM . CCM1 has been reported to be responsible for
[54]
localization of ICAP1α to the nucleus, through the use of the N-terminus NLS . It was shown that only in
[42]
the presence on intact CCM1- ICAP1α interaction and functional NLS in CCM1 does ICAP1α localize to the
[47]
nucleus . However, ICAP1α also contains a NLS and drives CCM1 localization to the nucleus. In the absence
of ICAP1α, CCM1 is evenly spread throughout the cell, but in the presence ICAP1α, the CCM1 localizes to
the nucleus. Alanine walking in NLS1, NLS2, and NES in CCM1 showed that only NLS1 affected CCM1
localization. NLS2 and NES mutation showed identical CCM1 localization to wild-type CCM1. However,
functional ICAP1α is able to translocate NLS1 mutated CCM1 into the nucleus. However, the N-terminus of
CCM1 is unable to translocate ICAP1α with a deficient NLS. The addition of CCM1 to ICAP1α -silenced cells
results in CCM1 accumulation only in the cytoplasm, suggesting that ICAP1α drives localization of CCM1,
not the other way around .
[119]

CCM proteins modulate VEGF signaling
Mutations in CCM1 and CCM3 were found to increase translocation of β-catenin from the cytosol to
the nucleus. This leads to increased expression of various proteins such as VEGF-A, which can be further
reversed by the addition of a β catenin transcription inhibitor. The increased level of VEGR-A activates
VEGFR2, which is shown by increased VEGFR2 phosphorylation. This increases the endothelial cell (EC)
permeability, leading to vascular leakage. This phenotype can be rescued with the addition of VEGFR2
inhibitors in both in vitro and in vivo conditions. Interestingly, VEGF inhibition blocked the formation of
stress fiber formation. Therefore, the stress fiber formation is caused to some extent by VEGF signaling.
Furthermore, enhanced VEGF signaling results in increased cellular migration. A wound-healing assay
showed that CCM1 deficient cells had a 25% increase in migration compared to the cultured cells treated
with VEGF. However, this phenotype can be reversed in the CCM1-null cells with the treatment of VEGF
inhibitors. VEGFR2 phosphorylation results in downstream phosphorylation of β-catenin and VE-cadherin,
which results in disruption of interaction with α-catenin and p120 catenin respectively. This results in
[120]
[121]
translocation of β-catenin into the nucleus for further downstream effects. However, VEGF inhibitors were
not sufficient to inhibit the β-catenin and VE-cadherin disassociation seen in CCM1 deficiency, suggesting
that other mechanisms are involved for the disassociation .
[122]
CCM2 modulates MAPK signaling
CCM2 leads to downstream activation of p38 MAPK, which is upregulated in osmotic shock. It is still unclear
for the role of the CCM2 in the p38 MAPK activation pathway. One report stated that CCM2 localizes to the
cell membrane where it facilitates binding MEKK3 and RAC1 leading to activation of MAPK [17,22] . Another
report showed that CCM2 is able to bind to F-actin, suggesting that CCM2 forms a complex that links
RAC1-dependent actin reorganization to p38 MAPK signal pathway . Another report showed that the
[123]
signaling pathway is through phospholipase C (PLC). The complex of CCM2-RAC1 causes a change in PLC
cascade, leading to MAPK activation . While another report showed that CCM2 affects the JNK and MKK
[124]
signaling leading to an alternative pathway to promote MAPK activation .
[89]
CCM3 plays a role in Notch signaling
In recent years, several mechanisms for the pathogenesis of CCMs have been proposed such as decreased
Notch signaling , increased VEGF signaling , or increased ERK activity in the deficiency of CCM3 .
[127]
[125]
[126]
It has been recently reported that CCM3 affects EC function by regulation of DLL4 . Down-regulation
[128]
of CCM3 resulted in decreased expression of DLL4 and Notch4, but no change was observed in Notch1.
This was shown in both cell lines and brain tissue in CCM patients. In fact, the vascular phenotype found
in CCM3 mutants can be replicated through mutations in DLL4. Aberrant DLL4/notch signaling results in