Limits...
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.

Show MeSH

Related in: MedlinePlus

Mitochondrial dynamics is altered in STHdhQ111/Q111 cells.A, Mitochondria-targeted GFP (mitoGFP), was transfected in the striatal cells to visualize the mitochondria and cells were then assigned to one of four groups based on the morphology of their mitochondria (tubular, mixed, fragmented or swollen). STHdhQ111/Q111 cells had more fragmented mitochondria and less tubular mitochondria compared to STHdhQ7/Q7 cells, while mixed and swollen mitochondria were at similar levels in the two cell types. More than 200 cells of each cell type were assessed for each independent experiment. n = 3 B, Immunoblots revealed that STHdhQ111/Q111 cells exhibit reduced levels of Drp1, its active form which is phosphorylated at Ser616, and Opa1 compared to STHdhQ7/Q7 cells, whereas Mfn2 appears to be similar in both cell types. C, Quantitative analysis of immunoblots shows a significant reduction in Drp1, phosphorylated Drp1, and Opa1. n = 4. Data shown are mean ± SE. **P<0.01, ***P<0.001 vs. STHdhQ7/Q7.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3585875&req=5

pone-0057932-g001: Mitochondrial dynamics is altered in STHdhQ111/Q111 cells.A, Mitochondria-targeted GFP (mitoGFP), was transfected in the striatal cells to visualize the mitochondria and cells were then assigned to one of four groups based on the morphology of their mitochondria (tubular, mixed, fragmented or swollen). STHdhQ111/Q111 cells had more fragmented mitochondria and less tubular mitochondria compared to STHdhQ7/Q7 cells, while mixed and swollen mitochondria were at similar levels in the two cell types. More than 200 cells of each cell type were assessed for each independent experiment. n = 3 B, Immunoblots revealed that STHdhQ111/Q111 cells exhibit reduced levels of Drp1, its active form which is phosphorylated at Ser616, and Opa1 compared to STHdhQ7/Q7 cells, whereas Mfn2 appears to be similar in both cell types. C, Quantitative analysis of immunoblots shows a significant reduction in Drp1, phosphorylated Drp1, and Opa1. n = 4. Data shown are mean ± SE. **P<0.01, ***P<0.001 vs. STHdhQ7/Q7.

Mentions: Mitochondrial morphology is an important component of cellular metabolism and pathogenic processes. Mitochondria are dynamic and display a range of morphologies. Maintaining the appropriate proportion of each mitochondrial morphological state is essential for appropriate functioning of the cell. To gain insight into mitochondrial dynamics in mHtt expressing cells, we transfected mitochondria-targeted GFP into STHdhQ7/Q7 and STHdhQ111/Q111 cells. Cells were divided into four groups based on the morphology of the mitochondria: tubular, fragmented, mixed, and swollen as previously described [35]. In most cells of both genotypes the mitochondria (about 60%) exhibited mixed forms with predominantly short tubular shapes. Approximately 20% of STHdhQ7/Q7 cells had predominantly tubular mitochondria while less than 10% of STHdhQ111/Q111 cells showed this mitochondrial morphology. Interestingly, more than 30% of the STHdhQ111/Q111 cells presented with fragmented mitochondria which was significantly greater than the STHdhQ7/Q7 cells. Both cell types exhibited very few cells with swollen mitochondria (Fig. 1A). Similarly, a significant increase in fragmented neuronal mitochondria in the striatum of R6/2 mice was also observed (Figure S1). Next, we examined whether there was a difference in the expression of mitochondrial fusion or fission regulators between the two cell types. Immunoblot analyses revealed that Drp1 levels were significantly lower in STHdhQ111/Q111 cells than in STHdhQ7/Q7 cells. In addition Drp1 phosphorylated at serine 616, which is a more active form [36], [37], was also lower in STHdhQ111/Q111 cells compared to SThdhQ7/Q7 cells. Of interest, Opa1 expression was also significantly decreased in STHdhQ111/Q111 cells, while Mfn2 appears to be at a similar level between the two cell types (Fig. 1, B and C). Together these results indicate that proteins involved in the dynamics of mitochondrial fission/fusion are altered in STHdhQ111/Q111 cells.


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)

Mitochondrial dynamics is altered in STHdhQ111/Q111 cells.A, Mitochondria-targeted GFP (mitoGFP), was transfected in the striatal cells to visualize the mitochondria and cells were then assigned to one of four groups based on the morphology of their mitochondria (tubular, mixed, fragmented or swollen). STHdhQ111/Q111 cells had more fragmented mitochondria and less tubular mitochondria compared to STHdhQ7/Q7 cells, while mixed and swollen mitochondria were at similar levels in the two cell types. More than 200 cells of each cell type were assessed for each independent experiment. n = 3 B, Immunoblots revealed that STHdhQ111/Q111 cells exhibit reduced levels of Drp1, its active form which is phosphorylated at Ser616, and Opa1 compared to STHdhQ7/Q7 cells, whereas Mfn2 appears to be similar in both cell types. C, Quantitative analysis of immunoblots shows a significant reduction in Drp1, phosphorylated Drp1, and Opa1. n = 4. Data shown are mean ± SE. **P<0.01, ***P<0.001 vs. STHdhQ7/Q7.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3585875&req=5

pone-0057932-g001: Mitochondrial dynamics is altered in STHdhQ111/Q111 cells.A, Mitochondria-targeted GFP (mitoGFP), was transfected in the striatal cells to visualize the mitochondria and cells were then assigned to one of four groups based on the morphology of their mitochondria (tubular, mixed, fragmented or swollen). STHdhQ111/Q111 cells had more fragmented mitochondria and less tubular mitochondria compared to STHdhQ7/Q7 cells, while mixed and swollen mitochondria were at similar levels in the two cell types. More than 200 cells of each cell type were assessed for each independent experiment. n = 3 B, Immunoblots revealed that STHdhQ111/Q111 cells exhibit reduced levels of Drp1, its active form which is phosphorylated at Ser616, and Opa1 compared to STHdhQ7/Q7 cells, whereas Mfn2 appears to be similar in both cell types. C, Quantitative analysis of immunoblots shows a significant reduction in Drp1, phosphorylated Drp1, and Opa1. n = 4. Data shown are mean ± SE. **P<0.01, ***P<0.001 vs. STHdhQ7/Q7.
Mentions: Mitochondrial morphology is an important component of cellular metabolism and pathogenic processes. Mitochondria are dynamic and display a range of morphologies. Maintaining the appropriate proportion of each mitochondrial morphological state is essential for appropriate functioning of the cell. To gain insight into mitochondrial dynamics in mHtt expressing cells, we transfected mitochondria-targeted GFP into STHdhQ7/Q7 and STHdhQ111/Q111 cells. Cells were divided into four groups based on the morphology of the mitochondria: tubular, fragmented, mixed, and swollen as previously described [35]. In most cells of both genotypes the mitochondria (about 60%) exhibited mixed forms with predominantly short tubular shapes. Approximately 20% of STHdhQ7/Q7 cells had predominantly tubular mitochondria while less than 10% of STHdhQ111/Q111 cells showed this mitochondrial morphology. Interestingly, more than 30% of the STHdhQ111/Q111 cells presented with fragmented mitochondria which was significantly greater than the STHdhQ7/Q7 cells. Both cell types exhibited very few cells with swollen mitochondria (Fig. 1A). Similarly, a significant increase in fragmented neuronal mitochondria in the striatum of R6/2 mice was also observed (Figure S1). Next, we examined whether there was a difference in the expression of mitochondrial fusion or fission regulators between the two cell types. Immunoblot analyses revealed that Drp1 levels were significantly lower in STHdhQ111/Q111 cells than in STHdhQ7/Q7 cells. In addition Drp1 phosphorylated at serine 616, which is a more active form [36], [37], was also lower in STHdhQ111/Q111 cells compared to SThdhQ7/Q7 cells. Of interest, Opa1 expression was also significantly decreased in STHdhQ111/Q111 cells, while Mfn2 appears to be at a similar level between the two cell types (Fig. 1, B and C). Together these results indicate that proteins involved in the dynamics of mitochondrial fission/fusion are altered in STHdhQ111/Q111 cells.

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