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Lazarillo-related Lipocalins confer long-term protection against type I Spinocerebellar Ataxia degeneration contributing to optimize selective autophagy.

del Caño-Espinel M, Acebes JR, Sanchez D, Ganfornina MD - Mol Neurodegener (2015)

Bottom Line: GLaz beneficial effects persist throughout aging, and appears when expressed by degenerating neurons or by retinal support and glial cells.GLaz gain-of-function reduces cell death and the extent of ubiquitinated proteins accumulation, and decreases the expression of Atg8a/LC3, p62 mRNA and protein levels, and GstS1 induction.Down-regulation of selective autophagy causes similar and non-additive rescuing effects.

View Article: PubMed Central - PubMed

Affiliation: Instituto de Biología y Genética Molecular-Departamento de Bioquímica y Biología Molecular y Fisiología, Universidad de Valladolid-CSIC, c/ Sanz y Forés 3, 47003, Valladolid, Spain. manuela@ibgm.uva.es.

ABSTRACT

Background: A diverse set of neurodegenerative disorders are caused by abnormal extensions of polyglutamine (poly-Q) stretches in various, functionally unrelated proteins. A common feature of these diseases is altered proteostasis. Autophagy induction is part of the endogenous response to poly-Q protein expression. However, if autophagy is not resolved properly, clearance of toxic proteins or aggregates cannot occur effectively. Likewise, excessive autophagy induction can cause autophagic stress and neurodegeneration. The Lipocalins ApoD, Glial Lazarillo (GLaz) and Neural Lazarillo (NLaz) are neuroprotectors upon oxidative stress or aging. In this work we test whether these Lipocalins also protect against poly-Q-triggered deterioration of protein quality control systems.

Results: Using a Drosophila retinal degeneration model of Type-1 Spinocerebellar Ataxia (SCA1) combined with genetic manipulation of NLaz and GLaz expression, we demonstrate that both Lipocalins protect against SCA1 neurodegeneration. They are part of the endogenous transcriptional response to SCA1, and their effect is non-additive, suggesting participation in a similar mechanism. GLaz beneficial effects persist throughout aging, and appears when expressed by degenerating neurons or by retinal support and glial cells. GLaz gain-of-function reduces cell death and the extent of ubiquitinated proteins accumulation, and decreases the expression of Atg8a/LC3, p62 mRNA and protein levels, and GstS1 induction. Over-expression of GLaz is able to reduce p62 and ubiquitinated proteins levels when rapamycin-dependent and SCA1-dependent inductions of autophagy are combined. In the absence of neurodegeneration, GLaz loss-of-function increases Atg8a/LC3 mRNA and p62 protein levels without altering p62 mRNA levels. Knocking-down autophagy, by interfering with Atg8a or p62 expression or by expressing dominant-negative Atg1/ULK1 or Atg4a transgenes, rescues SCA1-dependent neurodegeneration in a similar extent to the protective effect of GLaz. Further GLaz-dependent improvement is concealed.

Conclusions: This work shows for the first time that a Lipocalin rescues neurons from pathogenic SCA1 degeneration by optimizing clearance of aggregation-prone proteins. GLaz modulates key autophagy genes and lipid-peroxide clearance responsive genes. Down-regulation of selective autophagy causes similar and non-additive rescuing effects. These data suggest that SCA1 neurodegeneration concurs with autophagic stress, and places Lazarillo-related Lipocalins as valuable players in the endogenous protection against the two major contributors to aging and neurodegeneration: ROS-dependent damage and proteostasis deterioration.

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Decrease of ubiquitinated proteins build-up attained by GLaz over-expression. A, Ubiquitin HRP-immunohistochemistry on paraffin sections of control, degenerated and GLaz-rescued retinas shows the decrease in ubiquitin labeling when over-expressing GLaz. B, Ubiquitinated protein load measured by ubiquitin immunoblot signal in different lines expressing the pathogenic hATXN182Q alone or in combination with GLaz-expressing transgenes. Statistical differences were assayed by Student’s t-test. *P < 0.05.
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Fig5: Decrease of ubiquitinated proteins build-up attained by GLaz over-expression. A, Ubiquitin HRP-immunohistochemistry on paraffin sections of control, degenerated and GLaz-rescued retinas shows the decrease in ubiquitin labeling when over-expressing GLaz. B, Ubiquitinated protein load measured by ubiquitin immunoblot signal in different lines expressing the pathogenic hATXN182Q alone or in combination with GLaz-expressing transgenes. Statistical differences were assayed by Student’s t-test. *P < 0.05.

Mentions: To address whether GLaz over-expression modifies any of the protein quality control systems altered in the SCA1fly model, we analyzed by immunohistochemistry and immunoblot the expression pattern of ubiquitinated proteins (Figure 5). Abundant ubiquitinated protein aggregates are evident upon neurodegeneration (Figure 5Ab,d) when compared to controls (driver alone, not shown, or GLaz over-expression in wild type background, Figure 5Aa). This accumulation is greatly reduced when GLaz is co-expressed with hATXN182Q (Figure 5Ac,e,f). A quantitative analysis by immunoblot demonstrates that over-expressing GLaz, either in the degenerating photoreceptor or in the native GLaz expression domain, results in a significant decrease of ubiquitinated protein levels (Figure 5B).Figure 5


Lazarillo-related Lipocalins confer long-term protection against type I Spinocerebellar Ataxia degeneration contributing to optimize selective autophagy.

del Caño-Espinel M, Acebes JR, Sanchez D, Ganfornina MD - Mol Neurodegener (2015)

Decrease of ubiquitinated proteins build-up attained by GLaz over-expression. A, Ubiquitin HRP-immunohistochemistry on paraffin sections of control, degenerated and GLaz-rescued retinas shows the decrease in ubiquitin labeling when over-expressing GLaz. B, Ubiquitinated protein load measured by ubiquitin immunoblot signal in different lines expressing the pathogenic hATXN182Q alone or in combination with GLaz-expressing transgenes. Statistical differences were assayed by Student’s t-test. *P < 0.05.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4374295&req=5

Fig5: Decrease of ubiquitinated proteins build-up attained by GLaz over-expression. A, Ubiquitin HRP-immunohistochemistry on paraffin sections of control, degenerated and GLaz-rescued retinas shows the decrease in ubiquitin labeling when over-expressing GLaz. B, Ubiquitinated protein load measured by ubiquitin immunoblot signal in different lines expressing the pathogenic hATXN182Q alone or in combination with GLaz-expressing transgenes. Statistical differences were assayed by Student’s t-test. *P < 0.05.
Mentions: To address whether GLaz over-expression modifies any of the protein quality control systems altered in the SCA1fly model, we analyzed by immunohistochemistry and immunoblot the expression pattern of ubiquitinated proteins (Figure 5). Abundant ubiquitinated protein aggregates are evident upon neurodegeneration (Figure 5Ab,d) when compared to controls (driver alone, not shown, or GLaz over-expression in wild type background, Figure 5Aa). This accumulation is greatly reduced when GLaz is co-expressed with hATXN182Q (Figure 5Ac,e,f). A quantitative analysis by immunoblot demonstrates that over-expressing GLaz, either in the degenerating photoreceptor or in the native GLaz expression domain, results in a significant decrease of ubiquitinated protein levels (Figure 5B).Figure 5

Bottom Line: GLaz beneficial effects persist throughout aging, and appears when expressed by degenerating neurons or by retinal support and glial cells.GLaz gain-of-function reduces cell death and the extent of ubiquitinated proteins accumulation, and decreases the expression of Atg8a/LC3, p62 mRNA and protein levels, and GstS1 induction.Down-regulation of selective autophagy causes similar and non-additive rescuing effects.

View Article: PubMed Central - PubMed

Affiliation: Instituto de Biología y Genética Molecular-Departamento de Bioquímica y Biología Molecular y Fisiología, Universidad de Valladolid-CSIC, c/ Sanz y Forés 3, 47003, Valladolid, Spain. manuela@ibgm.uva.es.

ABSTRACT

Background: A diverse set of neurodegenerative disorders are caused by abnormal extensions of polyglutamine (poly-Q) stretches in various, functionally unrelated proteins. A common feature of these diseases is altered proteostasis. Autophagy induction is part of the endogenous response to poly-Q protein expression. However, if autophagy is not resolved properly, clearance of toxic proteins or aggregates cannot occur effectively. Likewise, excessive autophagy induction can cause autophagic stress and neurodegeneration. The Lipocalins ApoD, Glial Lazarillo (GLaz) and Neural Lazarillo (NLaz) are neuroprotectors upon oxidative stress or aging. In this work we test whether these Lipocalins also protect against poly-Q-triggered deterioration of protein quality control systems.

Results: Using a Drosophila retinal degeneration model of Type-1 Spinocerebellar Ataxia (SCA1) combined with genetic manipulation of NLaz and GLaz expression, we demonstrate that both Lipocalins protect against SCA1 neurodegeneration. They are part of the endogenous transcriptional response to SCA1, and their effect is non-additive, suggesting participation in a similar mechanism. GLaz beneficial effects persist throughout aging, and appears when expressed by degenerating neurons or by retinal support and glial cells. GLaz gain-of-function reduces cell death and the extent of ubiquitinated proteins accumulation, and decreases the expression of Atg8a/LC3, p62 mRNA and protein levels, and GstS1 induction. Over-expression of GLaz is able to reduce p62 and ubiquitinated proteins levels when rapamycin-dependent and SCA1-dependent inductions of autophagy are combined. In the absence of neurodegeneration, GLaz loss-of-function increases Atg8a/LC3 mRNA and p62 protein levels without altering p62 mRNA levels. Knocking-down autophagy, by interfering with Atg8a or p62 expression or by expressing dominant-negative Atg1/ULK1 or Atg4a transgenes, rescues SCA1-dependent neurodegeneration in a similar extent to the protective effect of GLaz. Further GLaz-dependent improvement is concealed.

Conclusions: This work shows for the first time that a Lipocalin rescues neurons from pathogenic SCA1 degeneration by optimizing clearance of aggregation-prone proteins. GLaz modulates key autophagy genes and lipid-peroxide clearance responsive genes. Down-regulation of selective autophagy causes similar and non-additive rescuing effects. These data suggest that SCA1 neurodegeneration concurs with autophagic stress, and places Lazarillo-related Lipocalins as valuable players in the endogenous protection against the two major contributors to aging and neurodegeneration: ROS-dependent damage and proteostasis deterioration.

Show MeSH
Related in: MedlinePlus