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Name: Saroj Chakraborty

02/16/2016
Abnormal Intracellular Calcium Signaling and SNARE Dependent Exocytosis Contributes
to SOD1G93A Astrocyte-Mediated Toxicity in Amyotrophic Lateral Sclerosis.
Hibiki Kawamata et al.

Hypothesis and Relevance:


This manuscript hypothesizes that the abnormal intracellular calcium signaling and SNARE
dependent exocytosis contributes to SOD1G93A astrocyte mediated degeneration of motor
neurons in amyotrophic lateral sclerosis (ALS). The authors find out that the astrocytes from
SOD1 mutant or ALS patients can cause embryonic motor neurons damage or death in cultures.
So, they try to findout the mechanisms by which Astrocyte cause the damage to motor neurons.
The significance of the study is to demonstrate a novel proposed mechanisms in which
Intracellular Calcium signaling as well as Snare dependent vesicular release involved in
Astrocyte regulation that leads to ALS.

Summary
Figure 1:
In this figure, the researchers try to compare the intracellular calcium signaling in Wild type
(WT) cells and SOD1G93A+ (G93) mutant. They find significant difference in the level of
calcium signaling after providing ATP. They also make sure it is not influenced by any
mitochondrial calcium signal interfered or by the SERCA pump. But, it would be much more
clear if they showed the base line or without any reagent calcium response.
Figure 2:
Here, the authors show the mitochondrial involvement in intracellular calcium signaling, where
they find no significant involvement of mitochondria.
Figure 3:
The researchers examine the level of calcium load in the ER of mutant SOD1G93A, where they
find out that the increase calcium release from the ER is due to Calcium overload in ER, not
because of increased Er volume or Calreticulin expression. Again, they also find out higher ATP
production that may lead to increased vesicular release.
Figure 4:
The researchers show that the SOCE in SOD1G93A mutant astrocytes are abnormal and it is ER
calcium independent, they make sure the TRCA channel is close in that experiment and the
response they get is only SOCE and find out that in presence of inhibitor like 2-APB the cells
give SOCE, and concluded that the mutant cell format puncta without any signal from ER-

Calcium depletion. But, the experiment would be more clear if they showed the response after
blocking STIM1 or orai3 andmake sure it is only SOCE.
Figure 5:
The researchers examine for the role of glutathionylated STIM1 in abnormal SOCE in
SOD1G93A and find out a direct correlation between them. They also treat the cell BSO
(oxidant) and BHA (antioxidant) and find out a direct link in WT treated with BSO and
SOD1G93A treated with BHA.
Figure 6:
Here the researcher show that inhibition of exocytosis by dn SNARE SOD1G93A effect on
toxicity to motor neurons. And they find out a correlation of dnSNARE leads to lower toxicity.
Figure 7:
Here the researcher show that inhibition of exocytosis by dn SNARE SOD1G93A cause less
ESMN death in ACM treated astrocytes.
Figure 8:
Now, the researchers try to establish a direct correlation between inhibition of exocytosis and
onset of disease. And they find out a increasing number of motor neuron survival and lateonset
of disease in dnSNARE SOD1G93A cells.
Figure 9:
Here, they tested the disease progression is affected by dn SNARE SOD1G93A inhibition of
exocytosis. And find out that no significant change in that. This data is quite disappointing s they
try to establish a relation with the progression.
Assessment
The authors findings support that the intracellular calcium signaling as well as SOCE is altered
in astrocytes in SOD1G93A mutants. In addition to that, they tested the effect of exocytosis and
altered SOCE in disease onset and find out a direct correlation. Although, at the end they dont
find a relation in death of mouse in ALS. Overall, the research conducted is top class and able to
answer all the questions. . However, there are a lot of holes in the paper as well as generating
many questions and missing links, like the direct evidence of exocytosis and ALS, other
pathways of calcium entry may influence or not, may be it was an effect of ALS rather than a
cause etc. Overall, these data provides a strong starting point for many questions to be asked in
the future.