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Loss of anti-Bax function in Gerstmann-Sträussler-Scheinker syndrome-associated prion protein mutants.

Jodoin J, Misiewicz M, Makhijani P, Giannopoulos PN, Hammond J, Goodyer CG, LeBlanc AC - PLoS ONE (2009)

Bottom Line: However, except for the P102L(V), none of the mutants significantly inhibited Bax-mediated caspase activation.These results show that the cytosolic PrP generated from the GSS mutants is not as efficient as wild type PrP in inhibiting Bax-mediated cell death.Furthermore, these results indicate that the anti-Bax function is also disrupted in GSS-associated PrP mutants and is not associated with the difference between CJD and GSS.

View Article: PubMed Central - PubMed

Affiliation: Bloomfield Center for Research in Aging, Lady Davis Institute for Medical Research, Sir Mortimer B. Davis Jewish General Hospital, Montréal, Canada.

ABSTRACT
Previously, we have shown the loss of anti-Bax function in Creutzfeldt Jakob disease (CJD)-associated prion protein (PrP) mutants that are unable to generate cytosolic PrP (CyPrP). To determine if the anti-Bax function of PrP modulates the manifestation of prion diseases, we further investigated the anti-Bax function of eight familial Gerstmann-Sträussler-Scheinker Syndrome (GSS)-associated PrP mutants. These PrP mutants contained their respective methionine ((M)) or valine ((V)) at codon 129. All of the mutants lost their ability to prevent Bax-mediated chromatin condensation or DNA fragmentation in primary human neurons. In the breast carcinoma MCF-7 cells, the F198S(V), D202N(V), P102L(V) and Q217R(V) retained, whereas the P102L(M), P105L(V), Y145stop(M) and Q212P(M) PrP mutants lost their ability to inhibit Bax-mediated condensed chromatin. The inhibition of Bax-mediated condensed chromatin depended on the ability of the mutants to generate cytosolic PrP. However, except for the P102L(V), none of the mutants significantly inhibited Bax-mediated caspase activation. These results show that the cytosolic PrP generated from the GSS mutants is not as efficient as wild type PrP in inhibiting Bax-mediated cell death. Furthermore, these results indicate that the anti-Bax function is also disrupted in GSS-associated PrP mutants and is not associated with the difference between CJD and GSS.

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All GSS-associated PrP mutants lose their anti-Bax function in primary human neurons.A. Percentage of cell death assessed by condensed chromatin in human neurons transfected with pBud-EGFP or pBud-EGFP-Bax (control), pBud-EGFP/PrP or PrP mutants, and pBud-EGFP-Bax/PrP or PrP mutants carrying a methionine (M) or a valine (V) at codon 129. Data represent the mean±SEM of three independent experiments. At least 300 cells were counted for each condition. * indicates a statistically significant difference (p≤0.05) compared to the control. B. Percentage of apoptotic cells displaying DNA fragmentation assessed by TUNEL in human neurons transfected with pBud-EGFP-Bax (control), pBud-EGFP-Bax/PrP, or pBud-EGFP-Bax/PrP mutants. Data represent the mean±SEM of three independent experiments. At least 150 cells were counted for each condition. * indicates a statistically significant difference (p≤0.05) compared to the control.
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pone-0006647-g005: All GSS-associated PrP mutants lose their anti-Bax function in primary human neurons.A. Percentage of cell death assessed by condensed chromatin in human neurons transfected with pBud-EGFP or pBud-EGFP-Bax (control), pBud-EGFP/PrP or PrP mutants, and pBud-EGFP-Bax/PrP or PrP mutants carrying a methionine (M) or a valine (V) at codon 129. Data represent the mean±SEM of three independent experiments. At least 300 cells were counted for each condition. * indicates a statistically significant difference (p≤0.05) compared to the control. B. Percentage of apoptotic cells displaying DNA fragmentation assessed by TUNEL in human neurons transfected with pBud-EGFP-Bax (control), pBud-EGFP-Bax/PrP, or pBud-EGFP-Bax/PrP mutants. Data represent the mean±SEM of three independent experiments. At least 150 cells were counted for each condition. * indicates a statistically significant difference (p≤0.05) compared to the control.

Mentions: To determine the anti-Bax function of GSS-associated PrP mutants in a neuronal cell type, we assessed the ability of the GSS-associated mutants against Bax-mediated cell death in primary human neurons. None of the GSS-associated PrP mutants induced cell death assessed by condensed chromatin in primary human neurons (Fig. 5A). However, these mutants also did not protect against Bax-mediated condensed chromatin (Fig. 5A) or TUNEL positive staining (Fig. 5B) compared to WT PrP's. Because all mutants lost their ability to inhibit the downstream Bax-mediated events, these results inferred that the mutants were also unable to inhibit caspase activation. The cytosolic level of GSS PrP mutants could not be assessed in these cells because of the very low transfection efficiency and the high endogenous level of PrP in the human primary neurons. Nevertheless, these results show the loss of anti-Bax function in the GSS-associated PrP mutants in human primary neurons.


Loss of anti-Bax function in Gerstmann-Sträussler-Scheinker syndrome-associated prion protein mutants.

Jodoin J, Misiewicz M, Makhijani P, Giannopoulos PN, Hammond J, Goodyer CG, LeBlanc AC - PLoS ONE (2009)

All GSS-associated PrP mutants lose their anti-Bax function in primary human neurons.A. Percentage of cell death assessed by condensed chromatin in human neurons transfected with pBud-EGFP or pBud-EGFP-Bax (control), pBud-EGFP/PrP or PrP mutants, and pBud-EGFP-Bax/PrP or PrP mutants carrying a methionine (M) or a valine (V) at codon 129. Data represent the mean±SEM of three independent experiments. At least 300 cells were counted for each condition. * indicates a statistically significant difference (p≤0.05) compared to the control. B. Percentage of apoptotic cells displaying DNA fragmentation assessed by TUNEL in human neurons transfected with pBud-EGFP-Bax (control), pBud-EGFP-Bax/PrP, or pBud-EGFP-Bax/PrP mutants. Data represent the mean±SEM of three independent experiments. At least 150 cells were counted for each condition. * indicates a statistically significant difference (p≤0.05) compared to the control.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2722024&req=5

pone-0006647-g005: All GSS-associated PrP mutants lose their anti-Bax function in primary human neurons.A. Percentage of cell death assessed by condensed chromatin in human neurons transfected with pBud-EGFP or pBud-EGFP-Bax (control), pBud-EGFP/PrP or PrP mutants, and pBud-EGFP-Bax/PrP or PrP mutants carrying a methionine (M) or a valine (V) at codon 129. Data represent the mean±SEM of three independent experiments. At least 300 cells were counted for each condition. * indicates a statistically significant difference (p≤0.05) compared to the control. B. Percentage of apoptotic cells displaying DNA fragmentation assessed by TUNEL in human neurons transfected with pBud-EGFP-Bax (control), pBud-EGFP-Bax/PrP, or pBud-EGFP-Bax/PrP mutants. Data represent the mean±SEM of three independent experiments. At least 150 cells were counted for each condition. * indicates a statistically significant difference (p≤0.05) compared to the control.
Mentions: To determine the anti-Bax function of GSS-associated PrP mutants in a neuronal cell type, we assessed the ability of the GSS-associated mutants against Bax-mediated cell death in primary human neurons. None of the GSS-associated PrP mutants induced cell death assessed by condensed chromatin in primary human neurons (Fig. 5A). However, these mutants also did not protect against Bax-mediated condensed chromatin (Fig. 5A) or TUNEL positive staining (Fig. 5B) compared to WT PrP's. Because all mutants lost their ability to inhibit the downstream Bax-mediated events, these results inferred that the mutants were also unable to inhibit caspase activation. The cytosolic level of GSS PrP mutants could not be assessed in these cells because of the very low transfection efficiency and the high endogenous level of PrP in the human primary neurons. Nevertheless, these results show the loss of anti-Bax function in the GSS-associated PrP mutants in human primary neurons.

Bottom Line: However, except for the P102L(V), none of the mutants significantly inhibited Bax-mediated caspase activation.These results show that the cytosolic PrP generated from the GSS mutants is not as efficient as wild type PrP in inhibiting Bax-mediated cell death.Furthermore, these results indicate that the anti-Bax function is also disrupted in GSS-associated PrP mutants and is not associated with the difference between CJD and GSS.

View Article: PubMed Central - PubMed

Affiliation: Bloomfield Center for Research in Aging, Lady Davis Institute for Medical Research, Sir Mortimer B. Davis Jewish General Hospital, Montréal, Canada.

ABSTRACT
Previously, we have shown the loss of anti-Bax function in Creutzfeldt Jakob disease (CJD)-associated prion protein (PrP) mutants that are unable to generate cytosolic PrP (CyPrP). To determine if the anti-Bax function of PrP modulates the manifestation of prion diseases, we further investigated the anti-Bax function of eight familial Gerstmann-Sträussler-Scheinker Syndrome (GSS)-associated PrP mutants. These PrP mutants contained their respective methionine ((M)) or valine ((V)) at codon 129. All of the mutants lost their ability to prevent Bax-mediated chromatin condensation or DNA fragmentation in primary human neurons. In the breast carcinoma MCF-7 cells, the F198S(V), D202N(V), P102L(V) and Q217R(V) retained, whereas the P102L(M), P105L(V), Y145stop(M) and Q212P(M) PrP mutants lost their ability to inhibit Bax-mediated condensed chromatin. The inhibition of Bax-mediated condensed chromatin depended on the ability of the mutants to generate cytosolic PrP. However, except for the P102L(V), none of the mutants significantly inhibited Bax-mediated caspase activation. These results show that the cytosolic PrP generated from the GSS mutants is not as efficient as wild type PrP in inhibiting Bax-mediated cell death. Furthermore, these results indicate that the anti-Bax function is also disrupted in GSS-associated PrP mutants and is not associated with the difference between CJD and GSS.

Show MeSH
Related in: MedlinePlus