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FUS Interacts with HSP60 to Promote Mitochondrial Damage.

Deng J, Yang M, Chen Y, Chen X, Liu J, Sun S, Cheng H, Li Y, Bigio EH, Mesulam M, Xu Q, Du S, Fushimi K, Zhu L, Wu JY - PLoS Genet. (2015)

Bottom Line: Down-regulating HSP60 reduces mitochondrially localized FUS and partially rescues mitochondrial defects and neurodegenerative phenotypes caused by FUS expression in transgenic flies.This is the first report of direct mitochondrial targeting by a nuclear protein associated with neurodegeneration, suggesting that mitochondrial impairment may represent a critical event in different forms of FUS-proteinopathies and a common pathological feature for both ALS-FUS and FTLD-FUS.Our study offers a potential explanation for the highly heterogeneous nature and complex genetic presentation of different forms of FUS-proteinopathies.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Brain and Cognitive Science, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China; University of Chinese Academy of Sciences, Beijing, China.

ABSTRACT
FUS-proteinopathies, a group of heterogeneous disorders including ALS-FUS and FTLD-FUS, are characterized by the formation of inclusion bodies containing the nuclear protein FUS in the affected patients. However, the underlying molecular and cellular defects remain unclear. Here we provide evidence for mitochondrial localization of FUS and its induction of mitochondrial damage. Remarkably, FTLD-FUS brain samples show increased FUS expression and mitochondrial defects. Biochemical and genetic data demonstrate that FUS interacts with a mitochondrial chaperonin, HSP60, and that FUS translocation to mitochondria is, at least in part, mediated by HSP60. Down-regulating HSP60 reduces mitochondrially localized FUS and partially rescues mitochondrial defects and neurodegenerative phenotypes caused by FUS expression in transgenic flies. This is the first report of direct mitochondrial targeting by a nuclear protein associated with neurodegeneration, suggesting that mitochondrial impairment may represent a critical event in different forms of FUS-proteinopathies and a common pathological feature for both ALS-FUS and FTLD-FUS. Our study offers a potential explanation for the highly heterogeneous nature and complex genetic presentation of different forms of FUS-proteinopathies. Our data also suggest that mitochondrial damage may be a target in future development of diagnostic and therapeutic tools for FUS-proteinopathies, a group of devastating neurodegenerative diseases.

No MeSH data available.


Related in: MedlinePlus

The FUS protein is associated with mitochondria.(A)Highly purified mitochondria were prepared from the control or P525L-FUS-expressing stable HEK cell lines. The mitochondrial purity was confirmed by the detection of mitochondrial CoxIV and the absence of the cytoplasmic proteins such as GAPDH or nuclear protein Histone H3. The endogenous FUS or P525L-mutant FUS localized to mitochondria; and the P525L-mutant FUS showed increased levels of mitochondrial localization (lane 6), as compared with the endogenous FUS in mitochondria (lane 3). (B-D) IEM images of the control or FUS-expressing stable HEK cell lines show reduced mitochondrial sizes in cells overexpressing FUS. Arrows, FUS-immunostaining signals associated with mitochondria labeled with 10-nm immuno-gold particles; arrowheads, mitochondria showed damaged cristae with “onion-like” structure. Mitochondrial cristae in P525L-mutant FUS expressing cells were significantly more frequently disrupted than the control and Wt-FUS groups, with quantification shown in panel C. More than 50 mitochondria were quantified in each group, analyzed using Chi-square test (***:p<0.0001).(D) Quantification of mitochondrial size using Image J. Mitochondria in Wt- or P525L-mutant FUS expressing cells were significantly smaller than the control group. At least 50 mitochondria were quantified in each group, analyzed using one-way ANOVA with Bonferronipost test (***: p<0.0001). (E) Mitochondrion-associated FUS immunostaining signals were significantly increased in Wt or P525L FUS expressing cells as compared with the Ctr. At least 60 mitochondria were quantified in each group, analyzed using one-way ANOVA with Bonferronipost test (**: p<0.01; ***: p<0.0001).
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pgen.1005357.g003: The FUS protein is associated with mitochondria.(A)Highly purified mitochondria were prepared from the control or P525L-FUS-expressing stable HEK cell lines. The mitochondrial purity was confirmed by the detection of mitochondrial CoxIV and the absence of the cytoplasmic proteins such as GAPDH or nuclear protein Histone H3. The endogenous FUS or P525L-mutant FUS localized to mitochondria; and the P525L-mutant FUS showed increased levels of mitochondrial localization (lane 6), as compared with the endogenous FUS in mitochondria (lane 3). (B-D) IEM images of the control or FUS-expressing stable HEK cell lines show reduced mitochondrial sizes in cells overexpressing FUS. Arrows, FUS-immunostaining signals associated with mitochondria labeled with 10-nm immuno-gold particles; arrowheads, mitochondria showed damaged cristae with “onion-like” structure. Mitochondrial cristae in P525L-mutant FUS expressing cells were significantly more frequently disrupted than the control and Wt-FUS groups, with quantification shown in panel C. More than 50 mitochondria were quantified in each group, analyzed using Chi-square test (***:p<0.0001).(D) Quantification of mitochondrial size using Image J. Mitochondria in Wt- or P525L-mutant FUS expressing cells were significantly smaller than the control group. At least 50 mitochondria were quantified in each group, analyzed using one-way ANOVA with Bonferronipost test (***: p<0.0001). (E) Mitochondrion-associated FUS immunostaining signals were significantly increased in Wt or P525L FUS expressing cells as compared with the Ctr. At least 60 mitochondria were quantified in each group, analyzed using one-way ANOVA with Bonferronipost test (**: p<0.01; ***: p<0.0001).

Mentions: To examine the relationship between FUS and mitochondria, we purified mitochondria from the HEK293 cells expressing the vector control or the ALS-associatedP525L-mutant FUS following published protocols [53,54] (Fig 3). Western blotting experiments demonstrate that our mitochondrial preparations were highly enriched in mitochondrial proteins (such as CoxIV) but devoid of either cytoplasmic proteins (such as GAPDH) or nuclear proteins (e.g., Histone H3) (Fig 3A). The endogenous FUS protein was consistently detected in these highly purified mitochondrial preparations (lane3 in Fig 3A). Although the level of P525L-mutant FUS protein detected was higher than the endogenous Wt-FUS protein in both the purified mitochondria and cytosol (see Fig 3A, the upper band was the P525L-mutant FUS; the lower band, the endogenous FUS), the endogenous FUS protein was clearly detected in the purified mitochondria (Fig 3A, lane 3 and lane 6). These data demonstrate that both the endogenous wild-type and transfected ALS-mutant FUS are translocated to mitochondria.


FUS Interacts with HSP60 to Promote Mitochondrial Damage.

Deng J, Yang M, Chen Y, Chen X, Liu J, Sun S, Cheng H, Li Y, Bigio EH, Mesulam M, Xu Q, Du S, Fushimi K, Zhu L, Wu JY - PLoS Genet. (2015)

The FUS protein is associated with mitochondria.(A)Highly purified mitochondria were prepared from the control or P525L-FUS-expressing stable HEK cell lines. The mitochondrial purity was confirmed by the detection of mitochondrial CoxIV and the absence of the cytoplasmic proteins such as GAPDH or nuclear protein Histone H3. The endogenous FUS or P525L-mutant FUS localized to mitochondria; and the P525L-mutant FUS showed increased levels of mitochondrial localization (lane 6), as compared with the endogenous FUS in mitochondria (lane 3). (B-D) IEM images of the control or FUS-expressing stable HEK cell lines show reduced mitochondrial sizes in cells overexpressing FUS. Arrows, FUS-immunostaining signals associated with mitochondria labeled with 10-nm immuno-gold particles; arrowheads, mitochondria showed damaged cristae with “onion-like” structure. Mitochondrial cristae in P525L-mutant FUS expressing cells were significantly more frequently disrupted than the control and Wt-FUS groups, with quantification shown in panel C. More than 50 mitochondria were quantified in each group, analyzed using Chi-square test (***:p<0.0001).(D) Quantification of mitochondrial size using Image J. Mitochondria in Wt- or P525L-mutant FUS expressing cells were significantly smaller than the control group. At least 50 mitochondria were quantified in each group, analyzed using one-way ANOVA with Bonferronipost test (***: p<0.0001). (E) Mitochondrion-associated FUS immunostaining signals were significantly increased in Wt or P525L FUS expressing cells as compared with the Ctr. At least 60 mitochondria were quantified in each group, analyzed using one-way ANOVA with Bonferronipost test (**: p<0.01; ***: p<0.0001).
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4559378&req=5

pgen.1005357.g003: The FUS protein is associated with mitochondria.(A)Highly purified mitochondria were prepared from the control or P525L-FUS-expressing stable HEK cell lines. The mitochondrial purity was confirmed by the detection of mitochondrial CoxIV and the absence of the cytoplasmic proteins such as GAPDH or nuclear protein Histone H3. The endogenous FUS or P525L-mutant FUS localized to mitochondria; and the P525L-mutant FUS showed increased levels of mitochondrial localization (lane 6), as compared with the endogenous FUS in mitochondria (lane 3). (B-D) IEM images of the control or FUS-expressing stable HEK cell lines show reduced mitochondrial sizes in cells overexpressing FUS. Arrows, FUS-immunostaining signals associated with mitochondria labeled with 10-nm immuno-gold particles; arrowheads, mitochondria showed damaged cristae with “onion-like” structure. Mitochondrial cristae in P525L-mutant FUS expressing cells were significantly more frequently disrupted than the control and Wt-FUS groups, with quantification shown in panel C. More than 50 mitochondria were quantified in each group, analyzed using Chi-square test (***:p<0.0001).(D) Quantification of mitochondrial size using Image J. Mitochondria in Wt- or P525L-mutant FUS expressing cells were significantly smaller than the control group. At least 50 mitochondria were quantified in each group, analyzed using one-way ANOVA with Bonferronipost test (***: p<0.0001). (E) Mitochondrion-associated FUS immunostaining signals were significantly increased in Wt or P525L FUS expressing cells as compared with the Ctr. At least 60 mitochondria were quantified in each group, analyzed using one-way ANOVA with Bonferronipost test (**: p<0.01; ***: p<0.0001).
Mentions: To examine the relationship between FUS and mitochondria, we purified mitochondria from the HEK293 cells expressing the vector control or the ALS-associatedP525L-mutant FUS following published protocols [53,54] (Fig 3). Western blotting experiments demonstrate that our mitochondrial preparations were highly enriched in mitochondrial proteins (such as CoxIV) but devoid of either cytoplasmic proteins (such as GAPDH) or nuclear proteins (e.g., Histone H3) (Fig 3A). The endogenous FUS protein was consistently detected in these highly purified mitochondrial preparations (lane3 in Fig 3A). Although the level of P525L-mutant FUS protein detected was higher than the endogenous Wt-FUS protein in both the purified mitochondria and cytosol (see Fig 3A, the upper band was the P525L-mutant FUS; the lower band, the endogenous FUS), the endogenous FUS protein was clearly detected in the purified mitochondria (Fig 3A, lane 3 and lane 6). These data demonstrate that both the endogenous wild-type and transfected ALS-mutant FUS are translocated to mitochondria.

Bottom Line: Down-regulating HSP60 reduces mitochondrially localized FUS and partially rescues mitochondrial defects and neurodegenerative phenotypes caused by FUS expression in transgenic flies.This is the first report of direct mitochondrial targeting by a nuclear protein associated with neurodegeneration, suggesting that mitochondrial impairment may represent a critical event in different forms of FUS-proteinopathies and a common pathological feature for both ALS-FUS and FTLD-FUS.Our study offers a potential explanation for the highly heterogeneous nature and complex genetic presentation of different forms of FUS-proteinopathies.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Brain and Cognitive Science, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China; University of Chinese Academy of Sciences, Beijing, China.

ABSTRACT
FUS-proteinopathies, a group of heterogeneous disorders including ALS-FUS and FTLD-FUS, are characterized by the formation of inclusion bodies containing the nuclear protein FUS in the affected patients. However, the underlying molecular and cellular defects remain unclear. Here we provide evidence for mitochondrial localization of FUS and its induction of mitochondrial damage. Remarkably, FTLD-FUS brain samples show increased FUS expression and mitochondrial defects. Biochemical and genetic data demonstrate that FUS interacts with a mitochondrial chaperonin, HSP60, and that FUS translocation to mitochondria is, at least in part, mediated by HSP60. Down-regulating HSP60 reduces mitochondrially localized FUS and partially rescues mitochondrial defects and neurodegenerative phenotypes caused by FUS expression in transgenic flies. This is the first report of direct mitochondrial targeting by a nuclear protein associated with neurodegeneration, suggesting that mitochondrial impairment may represent a critical event in different forms of FUS-proteinopathies and a common pathological feature for both ALS-FUS and FTLD-FUS. Our study offers a potential explanation for the highly heterogeneous nature and complex genetic presentation of different forms of FUS-proteinopathies. Our data also suggest that mitochondrial damage may be a target in future development of diagnostic and therapeutic tools for FUS-proteinopathies, a group of devastating neurodegenerative diseases.

No MeSH data available.


Related in: MedlinePlus