A method for the analysis of the oligomerization profile of the Huntington’s disease-associated, aggregation-prone mutant huntingtin protein by isopycnic ultracentrifugation

Conformational diseases, such as Alzheimer’s, Parkinson’s and Huntington’s diseases as well as ataxias and fronto-temporal disorders, are part of common class of neurological disorders characterised by the aggregation and progressive accumulation of mutant proteins which display aberrant conformation. In particular, Huntington’s disease (HD) is caused by mutations leading to an abnormal expansion in the polyglutamine (poly-Q) tract of the huntingtin protein (HTT), leading to the formation of inclusion bodies in neurons of affected patients. Furthermore, recent experimental evidence is challenging the conventional view of the disease by revealing the ability of mutant HTT to be transferred between cells by means of extracellular vesicles (EVs), allowing the mutant protein to seed oligomers involving both the mutant and wild type forms of the protein. There is still no successful strategy to treat HD. In addition, the current understanding of the biological processes leading to the oligomerization and aggregation of proteins bearing the poly-Q tract has been derived from studies conducted on isolated poly-Q monomers and oligomers, whose structural properties are still unclear and often inconsistent. Here we describe a standardised biochemical approach to analyse by isopycnic ultracentrifugation the oligomerization of the N-terminal fragment of mutant HTT. The dynamic range of our method allows one to detect large and heterogeneous HTT complexes. Hence, it could be harnessed for the identification of novel molecular determinants responsible for the aggregation and the prion-like spreading properties of HTT in the context of HD. Equally, it provides a tool to test novel small molecules or bioactive compounds designed to inhibit the aggregation of mutant HTT

Influence of ewe feeding systems on meat quality of suckling lambs

In recent years interest has grown in the zootechnical exploitation of environmental feeding resources, above all in marginal areas. The survival of these areas is linked to the development of the limited available resources. Of these, natural pastures represent one of the most important, not only because their zootechnical utilisation permits savings in alimentary costs, but above all because it results in better quality dairy and meat products. The aim of this study is to verify if and to what level ewe feeding systems influence the meat quality of suckling lambs

Early onset effects of single substrate accumulation recapitulate major features of LSD in patient-derived lysosomes.

Lysosome functions mainly rely on their ability to either degrade substrates or release them into the extracellular space. Lysosomal storage disorders (LSDs) are commonly characterized by a chronic lysosomal accumulation of different substrates, thereby causing lysosomal dysfunctions and secretion defects. However, the early effects of substrate accumulation on lysosomal homeostasis have not been analyzed so far. Here, we describe how the acute accumulation of a single substrate determines a rapid centripetal redistribution of the lysosomes, triggering their expansion and reducing their secretion, by limiting the motility of these organelles toward the plasma membrane. Moreover, we provide evidence that such defects could be explained by a trapping mechanism exerted by the extensive contacts between the enlarged lysosomes and the highly intertwined membrane structures of the endoplasmic reticulum which might represent a crucial biological cue ultimately leading to the clinically relevant secondary defects observed in the LSD experimental models and patients

Competitive binding of extracellular accumulated heparan sulfate reduces lysosomal storage defects and triggers neuronal differentiation in a model of Mucopolysaccharidosis IIIB

Mucopolysaccharidoses (MPSs) are a group of inherited lysosomal storage disorders associated with the deficiency of lysosomal enzymes involved in glycosaminoglycan (GAG) degradation. The resulting cellular accumulation of GAGs is responsible for widespread tissue and organ dysfunctions. The MPS III, caused by mutations in the genes responsible for the degradation of heparan sulfate (HS), includes four subtypes (A, B, C, and D) that present significant neurological manifestations such as progressive cognitive decline and behavioral disorders. The established treatments for the MPS III do not cure the disease but only ameliorate non-neurological clinical symptoms. We previously demonstrated that the natural variant of the hepatocyte growth factor NK1 reduces the lysosomal pathology and reactivates impaired growth factor signaling in fibroblasts from MPS IIIB patients. Here, we show that the recombinant NK1 is effective in rescuing the morphological and functional dysfunctions of lysosomes in a neuronal cellular model of the MPS IIIB. More importantly, NK1 treatment is able to stimulate neuronal differentiation of neuroblastoma SK-NBE cells stable silenced for the NAGLU gene causative of the MPS IIIB. These results provide the basis for the development of a novel approach to possibly correct the neurological phenotypes of the MPS IIIB as well as of other MPSs characterized by the accumulation of HS and progressive neurodegeneration