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  • Calpastatin ablation aggravates the molecular phenotype in cell and animal models of Huntington disease.

Calpastatin ablation aggravates the molecular phenotype in cell and animal models of Huntington disease.

Neuropharmacology (2018-01-23)
Jonasz Jeremiasz Weber, Simon Johannes Kloock, Maike Nagel, Midea Malena Ortiz-Rios, Julian Hofmann, Olaf Riess, Huu Phuc Nguyen
ABSTRACT

Deciphering the molecular pathology of Huntington disease is of particular importance, not only for a better understanding of this neurodegenerative disease, but also to identify potential therapeutic targets. The polyglutamine-expanded disease protein huntingtin was shown to undergo proteolysis, which results in the accumulation of toxic and aggregation-prone fragments. Amongst several classes of proteolytic enzymes responsible for huntingtin processing, the group of calcium-activated calpains has been found to be a significant mediator of the disease protein toxicity. To confirm the impact of calpain-mediated huntingtin cleavage in Huntington disease, we analysed the effect of depleting or overexpressing the endogenous calpain inhibitor calpastatin in HEK293T cells transfected with wild-type or polyglutamine-expanded huntingtin. Moreover, we crossbred huntingtin knock-in mice with calpastatin knockout animals to assess its effect not only on huntingtin cleavage and aggregation but also additional molecular markers. We demonstrated that a reduced or ablated expression of calpastatin triggers calpain overactivation and a consequently increased mutant huntingtin cleavage in cells and in vivo. These alterations were accompanied by an elevated formation of predominantly cytoplasmic huntingtin aggregates. On the other hand, overexpression of calpastatin in cells attenuated huntingtin fragmentation and aggregation. In addition, we observed an enhanced cleavage of DARPP-32, p35 and synapsin-1 in neuronal tissue upon calpain overactivation. Our results corroborate the important role of calpains in the molecular pathogenesis of Huntington disease and endorse targeting these proteolytic enzymes as a therapeutic approach.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
MISSION® esiRNA, targeting RLUC
Sigma-Aldrich
MDL 28170, ≥90% (TLC)