Science and Tech

New clues about the origin of a rare disease

[Img #67883]

Limb-girdle muscular dystrophy type 3 (LGMD D3) is a rare disease that causes progressive muscle weakness, caused by point mutations in the hnRNPDL-2 protein. Belonging to the family of ribonucleoproteins (RNP), associated with RNA, it is a little-known protein with the ability to self-assemble to form functional amyloid structures. Amyloids are formed by the union of thousands of copies of the same protein to form very stable and structured fibers (protein aggregates). Their formation is frequently associated with diseases such as Parkinson’s and Alzheimer’s, but they are also used by different organisms for functional purposes, although the number of functional amyloids described in humans is still low.

Researchers from the Autonomous University of Barcelona (UAB) have determined the structure of the amyloid fibers formed by the hnRNPDL-2 protein. Their architecture and activity suggest that they are stable, nontoxic amyloid fibers that bind nucleic acids in their aggregated state. The results indicate that LGMD D3 could be a protein loss-of-function disease: the inability to form the amyloid structures described in the study would cause the pathology.

“Our study breaks with the hypothesis that the aggregation of this protein is the cause of the disease and proposes that it is the inability to form a fibrillar structure that has been selected by evolution to bind nucleic acids that would cause the pathology,” he indicates. Salvador Ventura, professor of Biochemistry and Molecular Biology and researcher at the Institute of Biotechnology and Biomedicine (IBB) of the UAB, who has directed the research together with the first author of the work, Javier Garcia-Pardo, researcher Juan de la Cierva-Incorporation of the IBB .

Javier Garcia-Pardo (left), Salvador Ventura and Andrea Bartolomé-Nafría, from the research team. (Photo: UAB)

Researchers have determined the structure of the amyloid fibers of the hnRNPDL-2 protein using high-resolution cryo-electron microscopy (cryo-EM). This is the first structure of a functional human amyloid made up of the complete protein to be resolved using this technique —previously it had only been done with structures made up of fragments of these proteins. It is also the first amyloid structure determined at high resolution by a Spanish group.

The structure of the protein differs from fibers of other pathological amyloid proteins by presenting a highly hydrophilic core, which includes the amino acid associated with LGMD D3. In this case, contrary to other diseases, the formation of amyloids is not toxic, but necessary for the function of the protein.

The results change the concept of the origin of the disease and how it should be treated, the researchers point out. “Previously, we thought, as occurs in many neurodegenerative diseases, that LGMD D3 originated because mutations in patients caused the initially soluble protein to form aggregates and, therefore, the search for antiaggregant molecules could be a potential therapy. Now we know that this would be a mistake, since it is the incorrect formation of the fiber that seems to trigger the disease; therefore, molecules that stabilize this structure or facilitate its formation would be the most appropriate”, points out Salvador Ventura.

Understanding the molecular structures of amyloids

Certain human amyloids can undergo both functional and pathologic aggregation, thus an understanding of their molecular structures is necessary to understand their distinctive qualities and functions. For example, RNPs similar to the one studied in this research, such as hnRNPA1 or FUS, are capable of forming functional amyloid fibers in response to cellular stress, but they can also harbor mutations responsible for diseases. These proteins are characterized by a modular architecture, which includes one or more nucleic acid-binding domains, together with disordered regions that are responsible for the formation of their assembly into functional or pathological amyloid structures.

“In recent years, the structures of various amyloid fibers made up of RNP fragments have been resolved. However, these assemblies may not necessarily coincide with those adopted in the context of complete proteins, as is the case with the structure obtained for hnRNPDL-2 resolved in our group”, explains Salvador Ventura. “In fact, our structure differs significantly from previous ones and questions some of the assumptions that were considered valid regarding the regulation of these proteins in cells,” he remarks.

Special techniques to resolve functional amyloids

To resolve the structure of hnRNPDL-2 in its assembled state, the team of researchers has used the cryo-EM technique, applying special techniques to resolve amyloid structures.

In the last two years, a significant number of amyloid fiber structures have been resolved with this technique, but these mainly correspond to pathological amyloids involved in systemic and neurodegenerative diseases.

“Our discovery demonstrates the power of cryo-EM to study the function of RNPs and the reasons for their link to disease. These proteins have been little studied so far, but they are associated with diseases such as Alzheimer’s, muscular dystrophies, cancer, neurodevelopmental disorders, and neuropsychiatric disorders. Thus, our objective now is to take advantage of the experience acquired in this technique to determine the fibrillar states of other functional amyloids and to study the effect of mutations, in order to better understand their implications for health and disease”, points out Salvador Ventura.

The development of this innovative technology at the UAB will allow researchers to exploit the recently installed cryo-EM platform at the Alba synchrotron, of which the University is a partner. Solving these types of structures requires a lot of computational power. The IBB Protein Folding and Conformational Diseases research group, led by Salvador Ventura, has just acquired a high-powered computer to carry out these calculations.

The work has been led by Salvador Ventura, with the participation of Javier García-Pardo, Andrea Bartolomé Nafría and Marcos Gil García, from the IBB research group. The structure has been resolved in collaboration with the group of Stefano Ricagno and Martino Bolognesi (Università degli Studi di Milano Statale and Unitech Nolimits Center).

The study is titled “Cryo-EM structure of hnRNPDL-2 fibrils, a functional amyloid associated with limb-girdle muscular dystrophy D3”. And it has been published in the academic journal Nature Communications. (Source: UAB. CC BY NC 4.0)

Source link