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Impaired mitochondrial dynamics and Nrf2 signaling contribute to compromised responses to oxidative stress in striatal cells expressing full-length mutant huntingtin.

Jin YN, Yu YV, Gundemir S, Jo C, Cui M, Tieu K, Johnson GV - PLoS ONE (2013)

Bottom Line: Oxidative stress leads to dramatic increases in the number of STHdh(Q111/Q111) cells containing swollen mitochondria, while STHdh(Q7/Q7) cells just show increases in the number of fragmented mitochondria. mHtt expression results in reduced activity of Nrf2, and activation of the Nrf2 pathway by the oxidant tBHQ is significantly impaired in STHdh(Q111/Q111) cells.Nrf2 expression does not differ between the two cell types, but STHdh(Q111/Q111) cells show reduced expression of Keap1 and p62, key modulators of Nrf2 signaling.These results suggest that mHtt disrupts Nrf2 signaling which contributes to impaired mitochondrial dynamics and may enhance susceptibility to oxidative stress in STHdh(Q111/Q111) cells.

View Article: PubMed Central - PubMed

Affiliation: Departmentsof Pharmacology and Physiology, University of Rochester, Rochester, New York, United States of America.

ABSTRACT
Huntington disease (HD) is an inherited neurodegenerative disease resulting from an abnormal expansion of polyglutamine in huntingtin (Htt). Compromised oxidative stress defense systems have emerged as a contributing factor to the pathogenesis of HD. Indeed activation of the Nrf2 pathway, which plays a prominent role in mediating antioxidant responses, has been considered as a therapeutic strategy for the treatment of HD. Given the fact that there is an interrelationship between impairments in mitochondrial dynamics and increased oxidative stress, in this present study we examined the effect of mutant Htt (mHtt) on these two parameters. STHdh(Q111/Q111) cells, striatal cells expressing mHtt, display more fragmented mitochondria compared to STHdh(Q7/Q7) cells, striatal cells expressing wild type Htt, concurrent with alterations in the expression levels of Drp1 and Opa1, key regulators of mitochondrial fission and fusion, respectively. Studies of mitochondrial dynamics using cell fusion and mitochondrial targeted photo-switchable Dendra revealed that mitochondrial fusion is significantly decreased in STHdh(Q111/Q111) cells. Oxidative stress leads to dramatic increases in the number of STHdh(Q111/Q111) cells containing swollen mitochondria, while STHdh(Q7/Q7) cells just show increases in the number of fragmented mitochondria. mHtt expression results in reduced activity of Nrf2, and activation of the Nrf2 pathway by the oxidant tBHQ is significantly impaired in STHdh(Q111/Q111) cells. Nrf2 expression does not differ between the two cell types, but STHdh(Q111/Q111) cells show reduced expression of Keap1 and p62, key modulators of Nrf2 signaling. In addition, STHdh(Q111/Q111) cells exhibit increases in autophagy, whereas the basal level of autophagy activation is low in STHdh(Q7/Q7) cells. These results suggest that mHtt disrupts Nrf2 signaling which contributes to impaired mitochondrial dynamics and may enhance susceptibility to oxidative stress in STHdh(Q111/Q111) cells.

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STHdhQ111/Q111 cells exhibit an increase in autophagy activity.A–B, The increase of LC3-I into LC3-II is a typical indicator of the autophagy. The total amount of LC3 including type I and II is significantly higher in STHdhQ7/Q7 cells than STHdhQ111/Q111 cells. In addition, the ratio of type II to type I seems to be greater in STHdhQ111/Q111 cells than in STHdhQ7/Q7 cells. n = 4. Data shown are mean ± SE. *P<0.05 vs. STHdhQ7/Q7. C, Cells were transfected with GFP-LC3 (microtubule-associated protein 1 light chain 3) and 24 h later GFP fluorescence images were monitored. LC3-I is a cytosolic form of LC3 and LC3-II is a membrane-bound form of LC3 that is implicated in autophagy. The localization of GFP-LC3 is the diffused cytosolic pattern in most of STHdhQ7/Q7 cells, while STHdhQ111/Q111 cells exhibit punctuate pattern of GFP-LC3 that indicates the active autophagic vesicles.
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pone-0057932-g009: STHdhQ111/Q111 cells exhibit an increase in autophagy activity.A–B, The increase of LC3-I into LC3-II is a typical indicator of the autophagy. The total amount of LC3 including type I and II is significantly higher in STHdhQ7/Q7 cells than STHdhQ111/Q111 cells. In addition, the ratio of type II to type I seems to be greater in STHdhQ111/Q111 cells than in STHdhQ7/Q7 cells. n = 4. Data shown are mean ± SE. *P<0.05 vs. STHdhQ7/Q7. C, Cells were transfected with GFP-LC3 (microtubule-associated protein 1 light chain 3) and 24 h later GFP fluorescence images were monitored. LC3-I is a cytosolic form of LC3 and LC3-II is a membrane-bound form of LC3 that is implicated in autophagy. The localization of GFP-LC3 is the diffused cytosolic pattern in most of STHdhQ7/Q7 cells, while STHdhQ111/Q111 cells exhibit punctuate pattern of GFP-LC3 that indicates the active autophagic vesicles.

Mentions: We next examined whether autophagy regulation is altered in STHdhQ111/Q111 cells compared to STHdhQ7/Q7 cells. First, we looked at the conversion of LC3-I into LC3-II as an indicator of autophagy activation. Although the total level of LC3 is significantly reduced in STHdhQ111/Q111 cells, the ratio of LC3-II to LC3-I appears to be increased in STHdhQ111/Q111 cells compared to STHdhQ7/Q7 cells (Fig. 9, A and B). To further assess the autophagy activation, we transfected GFP-LC3 into striatal cells and monitored the cellular distribution of GFP-LC3. As the activation of autophagy increases, GFP-LC3 relocates from the cytosol into autophagosome, resulting in the punctate pattern. We found that STHdhQ111/Q111 cells show numerous GFP-LC3 positive punctae, while STHdhQ7/Q7 cells showed only a few GFP-LC3 positive punctae (Fig. 9C). Together, these results indicate that STHdhQ111/Q111 cells exhibit highly activated autophagy signaling.


Impaired mitochondrial dynamics and Nrf2 signaling contribute to compromised responses to oxidative stress in striatal cells expressing full-length mutant huntingtin.

Jin YN, Yu YV, Gundemir S, Jo C, Cui M, Tieu K, Johnson GV - PLoS ONE (2013)

STHdhQ111/Q111 cells exhibit an increase in autophagy activity.A–B, The increase of LC3-I into LC3-II is a typical indicator of the autophagy. The total amount of LC3 including type I and II is significantly higher in STHdhQ7/Q7 cells than STHdhQ111/Q111 cells. In addition, the ratio of type II to type I seems to be greater in STHdhQ111/Q111 cells than in STHdhQ7/Q7 cells. n = 4. Data shown are mean ± SE. *P<0.05 vs. STHdhQ7/Q7. C, Cells were transfected with GFP-LC3 (microtubule-associated protein 1 light chain 3) and 24 h later GFP fluorescence images were monitored. LC3-I is a cytosolic form of LC3 and LC3-II is a membrane-bound form of LC3 that is implicated in autophagy. The localization of GFP-LC3 is the diffused cytosolic pattern in most of STHdhQ7/Q7 cells, while STHdhQ111/Q111 cells exhibit punctuate pattern of GFP-LC3 that indicates the active autophagic vesicles.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3585875&req=5

pone-0057932-g009: STHdhQ111/Q111 cells exhibit an increase in autophagy activity.A–B, The increase of LC3-I into LC3-II is a typical indicator of the autophagy. The total amount of LC3 including type I and II is significantly higher in STHdhQ7/Q7 cells than STHdhQ111/Q111 cells. In addition, the ratio of type II to type I seems to be greater in STHdhQ111/Q111 cells than in STHdhQ7/Q7 cells. n = 4. Data shown are mean ± SE. *P<0.05 vs. STHdhQ7/Q7. C, Cells were transfected with GFP-LC3 (microtubule-associated protein 1 light chain 3) and 24 h later GFP fluorescence images were monitored. LC3-I is a cytosolic form of LC3 and LC3-II is a membrane-bound form of LC3 that is implicated in autophagy. The localization of GFP-LC3 is the diffused cytosolic pattern in most of STHdhQ7/Q7 cells, while STHdhQ111/Q111 cells exhibit punctuate pattern of GFP-LC3 that indicates the active autophagic vesicles.
Mentions: We next examined whether autophagy regulation is altered in STHdhQ111/Q111 cells compared to STHdhQ7/Q7 cells. First, we looked at the conversion of LC3-I into LC3-II as an indicator of autophagy activation. Although the total level of LC3 is significantly reduced in STHdhQ111/Q111 cells, the ratio of LC3-II to LC3-I appears to be increased in STHdhQ111/Q111 cells compared to STHdhQ7/Q7 cells (Fig. 9, A and B). To further assess the autophagy activation, we transfected GFP-LC3 into striatal cells and monitored the cellular distribution of GFP-LC3. As the activation of autophagy increases, GFP-LC3 relocates from the cytosol into autophagosome, resulting in the punctate pattern. We found that STHdhQ111/Q111 cells show numerous GFP-LC3 positive punctae, while STHdhQ7/Q7 cells showed only a few GFP-LC3 positive punctae (Fig. 9C). Together, these results indicate that STHdhQ111/Q111 cells exhibit highly activated autophagy signaling.

Bottom Line: Oxidative stress leads to dramatic increases in the number of STHdh(Q111/Q111) cells containing swollen mitochondria, while STHdh(Q7/Q7) cells just show increases in the number of fragmented mitochondria. mHtt expression results in reduced activity of Nrf2, and activation of the Nrf2 pathway by the oxidant tBHQ is significantly impaired in STHdh(Q111/Q111) cells.Nrf2 expression does not differ between the two cell types, but STHdh(Q111/Q111) cells show reduced expression of Keap1 and p62, key modulators of Nrf2 signaling.These results suggest that mHtt disrupts Nrf2 signaling which contributes to impaired mitochondrial dynamics and may enhance susceptibility to oxidative stress in STHdh(Q111/Q111) cells.

View Article: PubMed Central - PubMed

Affiliation: Departmentsof Pharmacology and Physiology, University of Rochester, Rochester, New York, United States of America.

ABSTRACT
Huntington disease (HD) is an inherited neurodegenerative disease resulting from an abnormal expansion of polyglutamine in huntingtin (Htt). Compromised oxidative stress defense systems have emerged as a contributing factor to the pathogenesis of HD. Indeed activation of the Nrf2 pathway, which plays a prominent role in mediating antioxidant responses, has been considered as a therapeutic strategy for the treatment of HD. Given the fact that there is an interrelationship between impairments in mitochondrial dynamics and increased oxidative stress, in this present study we examined the effect of mutant Htt (mHtt) on these two parameters. STHdh(Q111/Q111) cells, striatal cells expressing mHtt, display more fragmented mitochondria compared to STHdh(Q7/Q7) cells, striatal cells expressing wild type Htt, concurrent with alterations in the expression levels of Drp1 and Opa1, key regulators of mitochondrial fission and fusion, respectively. Studies of mitochondrial dynamics using cell fusion and mitochondrial targeted photo-switchable Dendra revealed that mitochondrial fusion is significantly decreased in STHdh(Q111/Q111) cells. Oxidative stress leads to dramatic increases in the number of STHdh(Q111/Q111) cells containing swollen mitochondria, while STHdh(Q7/Q7) cells just show increases in the number of fragmented mitochondria. mHtt expression results in reduced activity of Nrf2, and activation of the Nrf2 pathway by the oxidant tBHQ is significantly impaired in STHdh(Q111/Q111) cells. Nrf2 expression does not differ between the two cell types, but STHdh(Q111/Q111) cells show reduced expression of Keap1 and p62, key modulators of Nrf2 signaling. In addition, STHdh(Q111/Q111) cells exhibit increases in autophagy, whereas the basal level of autophagy activation is low in STHdh(Q7/Q7) cells. These results suggest that mHtt disrupts Nrf2 signaling which contributes to impaired mitochondrial dynamics and may enhance susceptibility to oxidative stress in STHdh(Q111/Q111) cells.

Show MeSH
Related in: MedlinePlus