uu.seUppsala University Publications
Change search
ReferencesLink to record
Permanent link

Direct link
Prevention of HET-s prion aggregation by ribosomal RNA
(English)Manuscript (preprint) (Other academic)
Abstract [en]

The role of nucleic acids in prion aggregation / disaggregation has remained unclear.  Here, using HET-s prion from fungi Podospora anserina as a model system, we have studied the role of ribosomal RNA (rRNA) in its aggregation process in an unbiased manner. Our results show that rRNA, in particular domain V of rRNA from the large subunit of the ribosome, substantially prevents beta-amyloid aggregation of the HET-s prion in a concentration dependent manner. The sites of interaction of the HET-s prion on domain V rRNA have been identified, which overlap with the sites previously identified for the protein folding activity of the ribosome. This study provides a missing link for the role of rRNA based folding activity of the ribosome in the context of prion propagation.

National Category
Biochemistry and Molecular Biology
URN: urn:nbn:se:uu:diva-267736OAI: oai:DiVA.org:uu-267736DiVA: diva2:874132
Available from: 2015-11-25 Created: 2015-11-25 Last updated: 2016-01-13
In thesis
1. Protein Folding Activity of the Ribosome and Its Implication in Prion Processes
Open this publication in new window or tab >>Protein Folding Activity of the Ribosome and Its Implication in Prion Processes
2016 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

How the linear protein chains fold into their three-dimensional active conformation is one of the remaining puzzles of modern science. Other than molecular chaperones, ribosome - the cellular protein synthesis machinery, has also been implicated in protein folding. The active site of protein folding activity of the ribosome (PFAR) is in the domain V of the main RNA component of the large ribosomal subunit, which also constitutes the peptidyl transferase center.

We have characterized the mechanism of PFAR using ribosomes or ribosome-borne folding modulators (RFMs) and human carbonic anhydrase I (HCA) as a model system. RFMs from all three kingdoms of life showed PFAR.  By multiple addition of the denatured protein in the refolding assay we demonstrate that the RFMs can recycle efficiently to assist refolding of a new batch of denatured protein. The turnover of the RFMs, which includes release of the protein substrate, takes milliseconds. Furthermore, fast kinetics of HCA refolding suggests that an early folding intermediate is the substrate for PFAR. Our results demonstrate for the first time that PFAR is catalytic.

It was shown that two anti-prion drugs 6AP and GA specifically inhibit PFAR by binding to the domain V of the 23S / 25S rRNA. Using UV-crosslinking followed by primer extension we have identified the interaction sites of 6AP on domain V of 23S rRNA, which overlap with the protein binding sites, and are sensitive to mutagenesis. We find that 6AP and GA inhibit PFAR by direct competition with the substrate protein for the binding sites. Also, 6AP derivatives inhibit PFAR in the same order as their antiprion activity, 6AP8CF3 > 6AP8Cl > 6AP > 6APi. These results suggest involvement of PFAR in prion processes.

To clarify the role of PFAR in prion processes, we studied HET-s prion aggregation in the presence of domain V/ IV/II of rRNA. The rRNAs, especially domain V rRNA not only reduced HET-s aggregation, but also changed the morphology of the HET-s fibrils, which became shorter and less compact. These results show that PFAR actively prevents large amyloid aggregation and thus, possibly influence prion propagation. 

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2016. 53 p.
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology, ISSN 1651-6214 ; 1327
Ribosome, Protein folding, Prion disease, Antiprion drug, Competitive inhibition, PFAR, Amyloid
National Category
Biochemistry and Molecular Biology
Research subject
Biology with specialization in Molecular Biology
urn:nbn:se:uu:diva-267737 (URN)978-91-554-9429-2 (ISBN)
Public defence
2016-01-28, B22, Husargatan 3, Uppsala, 13:00 (English)
Available from: 2015-12-22 Created: 2015-11-25 Last updated: 2016-01-13

Open Access in DiVA

No full text

Biochemistry and Molecular Biology

Search outside of DiVA

GoogleGoogle Scholar
The number of downloads is the sum of all downloads of full texts. It may include eg previous versions that are now no longer available

Total: 72 hits
ReferencesLink to record
Permanent link

Direct link