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Cell line specific modulation of extracellular aβ42 by Hsp40.

Carnini A, Scott LO, Ahrendt E, Proft J, Winkfein RJ, Kim SW, Colicos MA, Braun JE - PLoS ONE (2012)

Bottom Line: Moreover, transient transfection of the stress induced Hsp40, decreased CAD levels of Aβ(42).Mutation of the conserved HPD motif within Hsp40 significantly reduced the Hsp40-mediated Aβ(42) increase in hippocampal cultures indicating the importance of this motif in regulating cellular Aβ(42).Therefore, increasing Hsp40 therapeutically with the intention of interfering with the pathogenic cascade leading to neurodegeneration in AD should be pursued with caution.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology and Pharmacology, Hotchkiss Brain Institute, University of Calgary, Calgary, Alberta, Canada.

ABSTRACT
Heat shock proteins (Hsps) are a set of molecular chaperones involved in cellular repair. They provide protective mechanisms that allow cells to survive potentially lethal insults, In response to a conditioning stress their expression is increased. Here we examined the connection between Hsps and Aβ(42), the amyloid peptide involved in the pathological sequence of Alzheimer's disease (AD). Extracellular Aβ(42) associates with neuronal cells and is a major constituent of senile plaques, one of the hallmarks of AD. Although Hsps are generally thought to prevent accumulation of misfolded proteins, there is a lack of mechanistic evidence that heat shock chaperones directly modulate Aβ(42) toxicity. In this study we show that neither extracellular Aβ(42) nor Aβ(42/)PrP(C) trigger the heat shock response in neurons. To address the influence of the neuroprotective heat shock response on cellular Aβ(42), Western analysis of Aβ(42) was performed following external Aβ(42) application. Five hours after a conditioning heat shock, Aβ(42) association with CAD cells was increased compared to control neurons. However, at forty-eight hours following heat shock Aβ(42) levels were reduced compared to that found for control cells. Moreover, transient transfection of the stress induced Hsp40, decreased CAD levels of Aβ(42). In contrast to CAD cells, hippocampal neurons transfected with Hsp40 retained Aβ(42) indicating that Hsp40 modulation of Aβ(42) proteostasis is cell specific. Mutation of the conserved HPD motif within Hsp40 significantly reduced the Hsp40-mediated Aβ(42) increase in hippocampal cultures indicating the importance of this motif in regulating cellular Aβ(42). Our data reveal a biochemical link between Hsp40 expression and Aβ(42) proteostasis that is cell specific. Therefore, increasing Hsp40 therapeutically with the intention of interfering with the pathogenic cascade leading to neurodegeneration in AD should be pursued with caution.

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Aβ42 or bovine rPrPC do not alter expression of the J proteins DnaJA1(Hdj2), DnaJA2(Rdj2), DnaJA3 (Tid1) or DnaJA4 in neural cells.CAD cells were incubated with 3 µM Aβ42 or 0.5 µM bovine rPrPC for 48 hours and washed in PBS prior to lysis. (A) Western analysis of heat shock proteins (B) The J proteins; DnaJA1(Hdj2), DnaJA2(Rdj2), DnaJA3 (Tid1) or DnaJA4 were detected by Western blot analysis. β-actin is shown as a loading control. Data are representative of 4 separate experiments.
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pone-0037755-g006: Aβ42 or bovine rPrPC do not alter expression of the J proteins DnaJA1(Hdj2), DnaJA2(Rdj2), DnaJA3 (Tid1) or DnaJA4 in neural cells.CAD cells were incubated with 3 µM Aβ42 or 0.5 µM bovine rPrPC for 48 hours and washed in PBS prior to lysis. (A) Western analysis of heat shock proteins (B) The J proteins; DnaJA1(Hdj2), DnaJA2(Rdj2), DnaJA3 (Tid1) or DnaJA4 were detected by Western blot analysis. β-actin is shown as a loading control. Data are representative of 4 separate experiments.

Mentions: Figure 5 shows that in the absence of cells, heat shock per se does not cause degradation or oligomerization of Aβ42. Also, PrPC does not initiate any shifts in the molecular weight of Aβ42 indicative of proteolysis or SDS-resistant oligomerization. In contrast a 15 kDa breakdown of PrPC was observed following heat shock. Figure 6 shows that neither PrPC nor Aβ42 were found to alter cellular levels of the constitutive chaperones DnaJA1/Hdj2, DnaJA2/Rdj2, DnaJA3/Tid1, DnaJA4, Hsc70 or the stress induced chaperones Hsp70, Hsp40 and Hsp25, indicating that a generalized reduction in these molecular chaperone levels is not an underlying mechanism of Aβ42 induced neurodegeneration.


Cell line specific modulation of extracellular aβ42 by Hsp40.

Carnini A, Scott LO, Ahrendt E, Proft J, Winkfein RJ, Kim SW, Colicos MA, Braun JE - PLoS ONE (2012)

Aβ42 or bovine rPrPC do not alter expression of the J proteins DnaJA1(Hdj2), DnaJA2(Rdj2), DnaJA3 (Tid1) or DnaJA4 in neural cells.CAD cells were incubated with 3 µM Aβ42 or 0.5 µM bovine rPrPC for 48 hours and washed in PBS prior to lysis. (A) Western analysis of heat shock proteins (B) The J proteins; DnaJA1(Hdj2), DnaJA2(Rdj2), DnaJA3 (Tid1) or DnaJA4 were detected by Western blot analysis. β-actin is shown as a loading control. Data are representative of 4 separate experiments.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3362613&req=5

pone-0037755-g006: Aβ42 or bovine rPrPC do not alter expression of the J proteins DnaJA1(Hdj2), DnaJA2(Rdj2), DnaJA3 (Tid1) or DnaJA4 in neural cells.CAD cells were incubated with 3 µM Aβ42 or 0.5 µM bovine rPrPC for 48 hours and washed in PBS prior to lysis. (A) Western analysis of heat shock proteins (B) The J proteins; DnaJA1(Hdj2), DnaJA2(Rdj2), DnaJA3 (Tid1) or DnaJA4 were detected by Western blot analysis. β-actin is shown as a loading control. Data are representative of 4 separate experiments.
Mentions: Figure 5 shows that in the absence of cells, heat shock per se does not cause degradation or oligomerization of Aβ42. Also, PrPC does not initiate any shifts in the molecular weight of Aβ42 indicative of proteolysis or SDS-resistant oligomerization. In contrast a 15 kDa breakdown of PrPC was observed following heat shock. Figure 6 shows that neither PrPC nor Aβ42 were found to alter cellular levels of the constitutive chaperones DnaJA1/Hdj2, DnaJA2/Rdj2, DnaJA3/Tid1, DnaJA4, Hsc70 or the stress induced chaperones Hsp70, Hsp40 and Hsp25, indicating that a generalized reduction in these molecular chaperone levels is not an underlying mechanism of Aβ42 induced neurodegeneration.

Bottom Line: Moreover, transient transfection of the stress induced Hsp40, decreased CAD levels of Aβ(42).Mutation of the conserved HPD motif within Hsp40 significantly reduced the Hsp40-mediated Aβ(42) increase in hippocampal cultures indicating the importance of this motif in regulating cellular Aβ(42).Therefore, increasing Hsp40 therapeutically with the intention of interfering with the pathogenic cascade leading to neurodegeneration in AD should be pursued with caution.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology and Pharmacology, Hotchkiss Brain Institute, University of Calgary, Calgary, Alberta, Canada.

ABSTRACT
Heat shock proteins (Hsps) are a set of molecular chaperones involved in cellular repair. They provide protective mechanisms that allow cells to survive potentially lethal insults, In response to a conditioning stress their expression is increased. Here we examined the connection between Hsps and Aβ(42), the amyloid peptide involved in the pathological sequence of Alzheimer's disease (AD). Extracellular Aβ(42) associates with neuronal cells and is a major constituent of senile plaques, one of the hallmarks of AD. Although Hsps are generally thought to prevent accumulation of misfolded proteins, there is a lack of mechanistic evidence that heat shock chaperones directly modulate Aβ(42) toxicity. In this study we show that neither extracellular Aβ(42) nor Aβ(42/)PrP(C) trigger the heat shock response in neurons. To address the influence of the neuroprotective heat shock response on cellular Aβ(42), Western analysis of Aβ(42) was performed following external Aβ(42) application. Five hours after a conditioning heat shock, Aβ(42) association with CAD cells was increased compared to control neurons. However, at forty-eight hours following heat shock Aβ(42) levels were reduced compared to that found for control cells. Moreover, transient transfection of the stress induced Hsp40, decreased CAD levels of Aβ(42). In contrast to CAD cells, hippocampal neurons transfected with Hsp40 retained Aβ(42) indicating that Hsp40 modulation of Aβ(42) proteostasis is cell specific. Mutation of the conserved HPD motif within Hsp40 significantly reduced the Hsp40-mediated Aβ(42) increase in hippocampal cultures indicating the importance of this motif in regulating cellular Aβ(42). Our data reveal a biochemical link between Hsp40 expression and Aβ(42) proteostasis that is cell specific. Therefore, increasing Hsp40 therapeutically with the intention of interfering with the pathogenic cascade leading to neurodegeneration in AD should be pursued with caution.

Show MeSH
Related in: MedlinePlus