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Kaya et al. Neuroimmunol Neuroinflammation 2019;6:5 I http://dx.doi.org/10.20517/2347-8659.2018.70 Page 9 of 15

reduction in cell viability and activation of the DNA damage response in SOD1 mutated cells was
established. Conversely, increased Speedy/RINGO expression enhanced cell viability and prevented the
DNA damage response in SOD1 mutated cells. These findings indicate that Speedy/RINGO confers a
similar protective benefit for both ALS motor neurons and degenerating primary hippocampal neurons. It
is therefore possible that Speedy/RINGO may someday play a role in treating neurodegenerative conditions
such as SCI.

Even though the exact mechanism for the protective role of Speedy/RINGO in p53-mediated apoptosis
requires further analysis, the effects are not due to the direct inhibition of calpain activity or p53 induction,
[13]
as calpain-mediated p53 induction was maintained even in the presence of Speedy/RINGO . The
mechanism may instead occur downstream from the p53 activation.

In addition to the aforementioned studies on degenerating primary neurons and ALS motor neurons, results
of carcinogenic [16-18] and astrocytic [15,63] proliferation studies point out that, among the downstream targets
of p53, ERK/MAPK and PI3K/AKT, signaling cascades are the most potent in terms of clarifying the p53-
dependent anti-apoptotic mechanism of Speedy/RINGO.

p53 regulation of ERK/MAPK and PI3K/AKT pathways in apoptosis
p53 is a tumor suppressive transcription factor that inhibits tumorigenesis under genotoxic conditions
by regulating gene expression. p53 induces anti-tumorigenesis mechanisms - including cell cycle arrest,
senescence and apoptosis - according to cellular conditions, type and intensity of stress signals [82-84] .
Similarly, the regulation of apoptosis, cell cycle arrest and proliferation by the ERK/MAPK pathway differs
depending on the type of stress signals received and the context of the cell [85,86] .

Under stress conditions, different signaling pathways can be triggered. For example, p38 [87-89] and ERK/
MAPK [90-93] activate p53 and induce its transcriptional activity. The reverse also occurs: p53 may activate
[96]
ERK/MAPK signaling [85,94,95] by inducing tyrosine kinase receptor DDR1 . In addition, p53 is also capable
of suppressing the ERK/MAPK pathway via activation of various phosphatases. These phosphatases then
dephosphorylate ERK and inhibit its anti-apoptotic function, leading to p53-dependent apoptosis [97,98] . It is
evident that interaction between p53 and ERK/MAPK signaling differs depending on cellular context and
the type of stress stimulus.

Another p53-related signaling pathway is PI3K/AKT. P13K/AKT has primarily been implicated in promoting
cell survival in response to extracellular signals [99-101] that regulate intracellular signaling cascade by
activating transmembrane receptors. This activation recruits PI3K isoforms to the plasma membrane which,
in turn, results in phosphorylation and activation of AKT. Activated AKT has a number of cell survival
stimulating effects through phosphorylation and inhibition of pro-apoptotic genes.

There is growing body of evidence indicating a negative regulatory function for p53 on cell survival in healthy
cells. In this mechanism, p53 binds to the promoter site of PTEN (a phosphatase and tensin homolog deleted
on chromosome ten) [102,103] . Active PTEN dephosphorylates the 3’ phosphate of phosphoinositol triphosphate
which results in the inhibition of the PI3K/AKT pathway. This inhibition subsequently causes a reduction in
phospho-AKT levels, which has been shown to cause G1 arrest in glioblastoma cells, but trigger apoptosis in
carcinomas [104,105] . In addition, p53 induction causes significant inhibition of PI3K/AKT in EB1 colon cancer
[94]
cells which strongly implies that inhibition of PI3K/AKT is essential for p53-dependent apoptosis .

These two pathways have been shown to be equally important for neuronal survival and regeneration after
nerve injury. Researchers found that 7 days after axotomy, ERK/MAPK and PI3K/AKT signaling activity was
[10]
increased, causing reduced apoptosis . Thus, it is reasonable to assume that the inhibitory effect of increased
p53 levels on the ERK/MAPK and PI3K/AKT pathways may be the major death signal for neurons in SCI.