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Neurodegeneration and functional impairments associated with glycogen synthase accumulation in a mouse model of Lafora disease.

Valles-Ortega J, Duran J, Garcia-Rocha M, Bosch C, Saez I, Pujadas L, Serafin A, Cañas X, Soriano E, Delgado-García JM, Gruart A, Guinovart JJ - EMBO Mol Med (2011)

Bottom Line: They also had LBs in the soma and some processes of PV(+) interneurons.This phenomenon was accompanied by the progressive loss of these neuronal cells and, importantly, neurophysiological alterations potentially related to impairment of hippocampal function.Our results emphasize the relevance of the laforin-malin complex in the control of glycogen metabolism and highlight altered glycogen accumulation as a key contributor to neurodegeneration in LD.

View Article: PubMed Central - PubMed

Affiliation: Institute for Research in Biomedicine (IRB Barcelona) Barcelona, Spain.

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Analysis of MGS in malin KO mouse brainsMGS protein is increased and accumulated in the insoluble fraction. Brain extracts from 11-month-old wild-type (WT) and malin knock-out (KO) mice were analysed. Total homogenates and the soluble and insoluble fractions resulting from low speed centrifugation were used for the biochemical analysis.A. Western blotting for MGS, GS phospho-serine 7 and 10 (pSer 7,10), GS phospho-serine 640 (pSer 640) and laforin. Actin was used as loading control.B,C. GS phoshorylation state. Densitometries from Western blot analysis are expressed as ratio of the signals from the enzyme phosphorylated at specific sites to total protein.C. Glycogen synthase (GS) activity measured in the presence of G6P.D. GS activity ratio (−G6P/+G6P). Data are expressed as mean ± SEM. *p < 0.05. ***p < 0.001. WT (n = 6), KO (n = 6).
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fig03: Analysis of MGS in malin KO mouse brainsMGS protein is increased and accumulated in the insoluble fraction. Brain extracts from 11-month-old wild-type (WT) and malin knock-out (KO) mice were analysed. Total homogenates and the soluble and insoluble fractions resulting from low speed centrifugation were used for the biochemical analysis.A. Western blotting for MGS, GS phospho-serine 7 and 10 (pSer 7,10), GS phospho-serine 640 (pSer 640) and laforin. Actin was used as loading control.B,C. GS phoshorylation state. Densitometries from Western blot analysis are expressed as ratio of the signals from the enzyme phosphorylated at specific sites to total protein.C. Glycogen synthase (GS) activity measured in the presence of G6P.D. GS activity ratio (−G6P/+G6P). Data are expressed as mean ± SEM. *p < 0.05. ***p < 0.001. WT (n = 6), KO (n = 6).

Mentions: Malin has been reported to be involved in the proteasomal clearance of laforin (Gentry et al, 2005) and MGS (Vilchez et al, 2007). Therefore, we analysed the MGS content and distribution in brain sections from WT and malin KO mice. For this purpose, we used 4- and 11-month-old animals to evaluate the progression with age. The polyglucosan inclusions were immunostained for MGS (Fig 1B), thereby indicating that LBs contain the GS protein and its catalytic product (Fig 1). Western blot analysis showed highly increased MGS in total homogenate from malin KO brains compared to controls. The levels of this protein were increased in the insoluble fraction, thus strengthening the results obtained from histochemistry. As polyglucosan inclusions apparently increased in number and size with age, we also analysed the levels of other glycogen-binding proteins as laforin, glycogenin—the priming enzyme for glycogen synthesis—and GP. The levels of laforin (Fig 3A), glycogen phosphorylase muscle (MGP) and brain (BGP) isoforms, and glycogenin (Supporting Information Fig 3) were also found to be increased and accumulated in the insoluble fraction. Interestingly, MGP levels were also increased in the soluble fraction where MGS, laforin, and BGP levels remained unchanged.


Neurodegeneration and functional impairments associated with glycogen synthase accumulation in a mouse model of Lafora disease.

Valles-Ortega J, Duran J, Garcia-Rocha M, Bosch C, Saez I, Pujadas L, Serafin A, Cañas X, Soriano E, Delgado-García JM, Gruart A, Guinovart JJ - EMBO Mol Med (2011)

Analysis of MGS in malin KO mouse brainsMGS protein is increased and accumulated in the insoluble fraction. Brain extracts from 11-month-old wild-type (WT) and malin knock-out (KO) mice were analysed. Total homogenates and the soluble and insoluble fractions resulting from low speed centrifugation were used for the biochemical analysis.A. Western blotting for MGS, GS phospho-serine 7 and 10 (pSer 7,10), GS phospho-serine 640 (pSer 640) and laforin. Actin was used as loading control.B,C. GS phoshorylation state. Densitometries from Western blot analysis are expressed as ratio of the signals from the enzyme phosphorylated at specific sites to total protein.C. Glycogen synthase (GS) activity measured in the presence of G6P.D. GS activity ratio (−G6P/+G6P). Data are expressed as mean ± SEM. *p < 0.05. ***p < 0.001. WT (n = 6), KO (n = 6).
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Related In: Results  -  Collection

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fig03: Analysis of MGS in malin KO mouse brainsMGS protein is increased and accumulated in the insoluble fraction. Brain extracts from 11-month-old wild-type (WT) and malin knock-out (KO) mice were analysed. Total homogenates and the soluble and insoluble fractions resulting from low speed centrifugation were used for the biochemical analysis.A. Western blotting for MGS, GS phospho-serine 7 and 10 (pSer 7,10), GS phospho-serine 640 (pSer 640) and laforin. Actin was used as loading control.B,C. GS phoshorylation state. Densitometries from Western blot analysis are expressed as ratio of the signals from the enzyme phosphorylated at specific sites to total protein.C. Glycogen synthase (GS) activity measured in the presence of G6P.D. GS activity ratio (−G6P/+G6P). Data are expressed as mean ± SEM. *p < 0.05. ***p < 0.001. WT (n = 6), KO (n = 6).
Mentions: Malin has been reported to be involved in the proteasomal clearance of laforin (Gentry et al, 2005) and MGS (Vilchez et al, 2007). Therefore, we analysed the MGS content and distribution in brain sections from WT and malin KO mice. For this purpose, we used 4- and 11-month-old animals to evaluate the progression with age. The polyglucosan inclusions were immunostained for MGS (Fig 1B), thereby indicating that LBs contain the GS protein and its catalytic product (Fig 1). Western blot analysis showed highly increased MGS in total homogenate from malin KO brains compared to controls. The levels of this protein were increased in the insoluble fraction, thus strengthening the results obtained from histochemistry. As polyglucosan inclusions apparently increased in number and size with age, we also analysed the levels of other glycogen-binding proteins as laforin, glycogenin—the priming enzyme for glycogen synthesis—and GP. The levels of laforin (Fig 3A), glycogen phosphorylase muscle (MGP) and brain (BGP) isoforms, and glycogenin (Supporting Information Fig 3) were also found to be increased and accumulated in the insoluble fraction. Interestingly, MGP levels were also increased in the soluble fraction where MGS, laforin, and BGP levels remained unchanged.

Bottom Line: They also had LBs in the soma and some processes of PV(+) interneurons.This phenomenon was accompanied by the progressive loss of these neuronal cells and, importantly, neurophysiological alterations potentially related to impairment of hippocampal function.Our results emphasize the relevance of the laforin-malin complex in the control of glycogen metabolism and highlight altered glycogen accumulation as a key contributor to neurodegeneration in LD.

View Article: PubMed Central - PubMed

Affiliation: Institute for Research in Biomedicine (IRB Barcelona) Barcelona, Spain.

Show MeSH
Related in: MedlinePlus