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Death Associated Protein Kinase (DAPK) -mediated neurodegenerative mechanisms in nematode excitotoxicity.

Del Rosario JS, Feldmann KG, Ahmed T, Amjad U, Ko B, An J, Mahmud T, Salama M, Mei S, Asemota D, Mano I - BMC Neurosci (2015)

Bottom Line: However, the molecular mechanism by which DAPK exerts its effect is controversial.We further show that some proposed mechanisms of DAPK's action (modulation of synaptic strength, involvement of the DANGER-related protein MAB-21, and autophagy) do not have a major role in nematode excitotoxicity.Our studies highlight the prominence of DAPK and Pin1/PINN-1 as conserved mediators of cell death processes in diverse scenarios of neurodegeneration.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, Pharmacology, and Neuroscience, Sophie Davis School of Biomedical Education (SBE), City College of New York (CCNY), The City University of New York (CUNY), New York, NY, USA. delrosariojohns@gmail.com.

ABSTRACT

Background: Excitotoxicity (the toxic overstimulation of neurons by the excitatory transmitter Glutamate) is a central process in widespread neurodegenerative conditions such as brain ischemia and chronic neurological diseases. Many mechanisms have been suggested to mediate excitotoxicity, but their significance across diverse excitotoxic scenarios remains unclear. Death Associated Protein Kinase (DAPK), a critical molecular switch that controls a range of key signaling and cell death pathways, has been suggested to have an important role in excitotoxicity. However, the molecular mechanism by which DAPK exerts its effect is controversial. A few distinct mechanisms have been suggested by single (sometimes contradicting) studies, and a larger array of potential mechanisms is implicated by the extensive interactome of DAPK.

Results: Here we analyze a well-characterized model of excitotoxicity in the nematode C. elegans to show that DAPK is an important mediator of excitotoxic neurodegeneration across a large evolutionary distance. We further show that some proposed mechanisms of DAPK's action (modulation of synaptic strength, involvement of the DANGER-related protein MAB-21, and autophagy) do not have a major role in nematode excitotoxicity. In contrast, Pin1/PINN-1 (a DAPK interaction-partner and a peptidyl-prolyl isomerase involved in chronic neurodegenerative conditions) suppresses neurodegeneration in our excitotoxicity model.

Conclusions: Our studies highlight the prominence of DAPK and Pin1/PINN-1 as conserved mediators of cell death processes in diverse scenarios of neurodegeneration.

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Related in: MedlinePlus

Treatments that block autophagy (and dramatically reduce neurodegeneration in other forms of necrotic neurodegeneration in C. elegans) have a reproducible but small effect in nematode excitotoxicity. A) A mutation in unc-51 shows a moderate effect on nematode excitotoxicity. *p < 0.05 ; ***p < 0.01 B) Treatment with the autophagy-blocking drug 3MA has a moderate effect on nematode excitotoxicity. ***p < 0.01 C) Independent repetition of the experiment shown in B with the addition of epistasis analysis. The data shows that the only reproducible effect of autophagy blockade is in L3. Although at this stage the combined effect of autophagy blockade and dapk-1 correlates with a model of independent action of these two factors, the moderate extent of effects limits the strength of such a conclusion.
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Fig4: Treatments that block autophagy (and dramatically reduce neurodegeneration in other forms of necrotic neurodegeneration in C. elegans) have a reproducible but small effect in nematode excitotoxicity. A) A mutation in unc-51 shows a moderate effect on nematode excitotoxicity. *p < 0.05 ; ***p < 0.01 B) Treatment with the autophagy-blocking drug 3MA has a moderate effect on nematode excitotoxicity. ***p < 0.01 C) Independent repetition of the experiment shown in B with the addition of epistasis analysis. The data shows that the only reproducible effect of autophagy blockade is in L3. Although at this stage the combined effect of autophagy blockade and dapk-1 correlates with a model of independent action of these two factors, the moderate extent of effects limits the strength of such a conclusion.

Mentions: To further study the possible involvement of autophagy in nematode excitotoxicity we examined the effect of inhibiting autophagy by genetic and chemical means. While a number of mutations and drugs have been used in the past, not all of them are available to us here. For example, bec-1 ko was previously used to monitor the requirement of this autophagy regulator for mec-4(d) –induced necrosis in early development [58]. However, the lethal effect of this mutation in later development, when most of the nematode excitotoxic necrosis occurs, prevents us from using this approach. Similarly, the vacuolar-type ATPase inhibitor bafilomycin is commonly used to block autophagy by elevating lysosomal pH [63]. However, the same V-ATPase is used in neurons to acidify synaptic vesicles as a means to provide the driving force for neurotransmitter loading [64-66], and therefore using bafilomycin can be expected to reduce neurotransmitter release. We therefore turned to use other means of intervention that are more compatible with our system: a mutation in the autophagy regulator unc-51 (using the e369 allele) and the chemical inhibitor 3MA (both used previously to show that degenerin-triggered neurodegeneration in C. elegans depends strongly on autophagy [58,60]). We noticed only a moderate effect for these two factors, evident in some developmental stages (Figure 4A and B). Such a moderate effect is in line with the reported effect of unc-51 on nuIs5 alone [60] (see discussion). We then used an independent set of experiments and epistasis analysis to determine if this moderate effect works independently of dapk-1 or in the same pathway. We noticed that the effect of blocking autophagy on the extent of excitotoxicity is reproducible only in one developmental stage (L3). Trying to determine if dapk-1 and 3MA work in the same pathway, we compared their observed combined effect (in the dapk-1 + 3MA combination) to the calculated expected effect if these two processes were completely independent. However, given the moderate size of the 3MA effect and the inherent variability in our experiments, it is currently difficult to determine if dapk-1 and autophagy work in the same pathway or independently. Nonetheless, the fact that the effect of autophagy is much more limited in size and duration than that of dapk-1 supported a continued search for other mechanisms by which dapk-1 might regulate excitotoxicity.Figure 4


Death Associated Protein Kinase (DAPK) -mediated neurodegenerative mechanisms in nematode excitotoxicity.

Del Rosario JS, Feldmann KG, Ahmed T, Amjad U, Ko B, An J, Mahmud T, Salama M, Mei S, Asemota D, Mano I - BMC Neurosci (2015)

Treatments that block autophagy (and dramatically reduce neurodegeneration in other forms of necrotic neurodegeneration in C. elegans) have a reproducible but small effect in nematode excitotoxicity. A) A mutation in unc-51 shows a moderate effect on nematode excitotoxicity. *p < 0.05 ; ***p < 0.01 B) Treatment with the autophagy-blocking drug 3MA has a moderate effect on nematode excitotoxicity. ***p < 0.01 C) Independent repetition of the experiment shown in B with the addition of epistasis analysis. The data shows that the only reproducible effect of autophagy blockade is in L3. Although at this stage the combined effect of autophagy blockade and dapk-1 correlates with a model of independent action of these two factors, the moderate extent of effects limits the strength of such a conclusion.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4414438&req=5

Fig4: Treatments that block autophagy (and dramatically reduce neurodegeneration in other forms of necrotic neurodegeneration in C. elegans) have a reproducible but small effect in nematode excitotoxicity. A) A mutation in unc-51 shows a moderate effect on nematode excitotoxicity. *p < 0.05 ; ***p < 0.01 B) Treatment with the autophagy-blocking drug 3MA has a moderate effect on nematode excitotoxicity. ***p < 0.01 C) Independent repetition of the experiment shown in B with the addition of epistasis analysis. The data shows that the only reproducible effect of autophagy blockade is in L3. Although at this stage the combined effect of autophagy blockade and dapk-1 correlates with a model of independent action of these two factors, the moderate extent of effects limits the strength of such a conclusion.
Mentions: To further study the possible involvement of autophagy in nematode excitotoxicity we examined the effect of inhibiting autophagy by genetic and chemical means. While a number of mutations and drugs have been used in the past, not all of them are available to us here. For example, bec-1 ko was previously used to monitor the requirement of this autophagy regulator for mec-4(d) –induced necrosis in early development [58]. However, the lethal effect of this mutation in later development, when most of the nematode excitotoxic necrosis occurs, prevents us from using this approach. Similarly, the vacuolar-type ATPase inhibitor bafilomycin is commonly used to block autophagy by elevating lysosomal pH [63]. However, the same V-ATPase is used in neurons to acidify synaptic vesicles as a means to provide the driving force for neurotransmitter loading [64-66], and therefore using bafilomycin can be expected to reduce neurotransmitter release. We therefore turned to use other means of intervention that are more compatible with our system: a mutation in the autophagy regulator unc-51 (using the e369 allele) and the chemical inhibitor 3MA (both used previously to show that degenerin-triggered neurodegeneration in C. elegans depends strongly on autophagy [58,60]). We noticed only a moderate effect for these two factors, evident in some developmental stages (Figure 4A and B). Such a moderate effect is in line with the reported effect of unc-51 on nuIs5 alone [60] (see discussion). We then used an independent set of experiments and epistasis analysis to determine if this moderate effect works independently of dapk-1 or in the same pathway. We noticed that the effect of blocking autophagy on the extent of excitotoxicity is reproducible only in one developmental stage (L3). Trying to determine if dapk-1 and 3MA work in the same pathway, we compared their observed combined effect (in the dapk-1 + 3MA combination) to the calculated expected effect if these two processes were completely independent. However, given the moderate size of the 3MA effect and the inherent variability in our experiments, it is currently difficult to determine if dapk-1 and autophagy work in the same pathway or independently. Nonetheless, the fact that the effect of autophagy is much more limited in size and duration than that of dapk-1 supported a continued search for other mechanisms by which dapk-1 might regulate excitotoxicity.Figure 4

Bottom Line: However, the molecular mechanism by which DAPK exerts its effect is controversial.We further show that some proposed mechanisms of DAPK's action (modulation of synaptic strength, involvement of the DANGER-related protein MAB-21, and autophagy) do not have a major role in nematode excitotoxicity.Our studies highlight the prominence of DAPK and Pin1/PINN-1 as conserved mediators of cell death processes in diverse scenarios of neurodegeneration.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, Pharmacology, and Neuroscience, Sophie Davis School of Biomedical Education (SBE), City College of New York (CCNY), The City University of New York (CUNY), New York, NY, USA. delrosariojohns@gmail.com.

ABSTRACT

Background: Excitotoxicity (the toxic overstimulation of neurons by the excitatory transmitter Glutamate) is a central process in widespread neurodegenerative conditions such as brain ischemia and chronic neurological diseases. Many mechanisms have been suggested to mediate excitotoxicity, but their significance across diverse excitotoxic scenarios remains unclear. Death Associated Protein Kinase (DAPK), a critical molecular switch that controls a range of key signaling and cell death pathways, has been suggested to have an important role in excitotoxicity. However, the molecular mechanism by which DAPK exerts its effect is controversial. A few distinct mechanisms have been suggested by single (sometimes contradicting) studies, and a larger array of potential mechanisms is implicated by the extensive interactome of DAPK.

Results: Here we analyze a well-characterized model of excitotoxicity in the nematode C. elegans to show that DAPK is an important mediator of excitotoxic neurodegeneration across a large evolutionary distance. We further show that some proposed mechanisms of DAPK's action (modulation of synaptic strength, involvement of the DANGER-related protein MAB-21, and autophagy) do not have a major role in nematode excitotoxicity. In contrast, Pin1/PINN-1 (a DAPK interaction-partner and a peptidyl-prolyl isomerase involved in chronic neurodegenerative conditions) suppresses neurodegeneration in our excitotoxicity model.

Conclusions: Our studies highlight the prominence of DAPK and Pin1/PINN-1 as conserved mediators of cell death processes in diverse scenarios of neurodegeneration.

Show MeSH
Related in: MedlinePlus