RNA modulates aggregation of the recombinant mammalian prion protein by direct interaction

Abstract

Recent studies have proposed that nucleic acids act as potential cofactors for protein aggregation and prionogenesis. By means of sedimentation, transmission electron microscopy, circular dichroism, static and dynamic light scattering, we have studied how RNA can influence the aggregation of the murine recombinant prion protein (rPrP). We find that RNA, independent of its sequence, source and size, modulates rPrP aggregation in a bimodal fashion, affecting both the extent and the rate of rPrP aggregation in a concentration dependent manner. Analogous to RNA-induced liquid-liquid phase transitions observed for other proteins implicated in neurodegenerative diseases, high protein to RNA ratios stimulate rPrP aggregation, while low ratios suppress it. However, the latter scenario also promotes formation of soluble oligomeric aggregates capable of seeding de novo rPrP aggregation. Furthermore, RNA co-aggregates with rPrP and thereby gains partial protection from RNase digestion. Our results also indicate that rPrP interacts with the RNAs with its N-terminus. In summary, this study elucidates the proposed adjuvant role of RNA in prion protein aggregation and propagation, and thus advocates an auxiliary role of the nucleic acids in protein aggregation in general.

Enzymatic degradation of RNA causes widespread protein aggregation in cell and tissue lysates

“…Such a stoichiometry suggests that each protein needs to be associated with one oligonucleotide to maintain solubility. These figures are in line with what has been observed for other aggregate-prone proteins such as the prion protein and FUS (Kovachev et al, 2019;Maharana et al, 2018). If seed-driven co-precipitation was a major force, a much lower oligo-to-protein ratio would be expected, as only the few seed proteins needs to be associated with the nucleic acid.…”

“…Recently, Maharana et al, showed that for several prion-like proteins, including FUS, these disparate effects can be explained by the ratio of protein to RNA; where excess of RNA promotes solubility and decreased amount of RNA induces phase-transition (Maharana et al, 2018). Similar results have also been observed for p53 and the prion protein (Kovachev et al, 2017;Kovachev et al, 2019).…”

“…Tau aggregation on the other hand has been reported to be stimulated by RNA (Kampers et al, 1996;Zhang et al, 2017), while the aggregation of Aβ has been reported to be inhibited (Mathura et al, 2005;Takahashi et al, 2009), a finding also confirmed here. Thus, it is clear that polyanions, at least in vitro, have diverse and sometimes paradoxical effects on protein structure and solubility, possibly depending on the protein / polyanion concentration (Kovachev et al, 2017;Kovachev et al, 2019;Maharana et al, 2018).…”

Liquid‐liquid phase separation and fibrillation of the prion protein modulated by a high‐affinity DNA aptamer

“…rPrP 90-231 at 10 µM in buffer without nucleic acids (black column), or with addition of increasing concentrations of A1 (shades of blue) or A2 (shades of green) or A1_mut (gray column). 5,62 Full-length PrP (PrP 23-231 ) contains at least three sequence motifs that bind NAs, 8 being able to form an interconnected polymer network with NAs. Quantification of rPrP 90-231 droplets number per 100 µm 2 area (bottom).…”

“…5,11 Furthermore, the observed NA effects on PrP aggregation are highly dependent on the PrP:NA binding stoichiometry. 5,62 Full-length PrP (PrP 23-231 ) contains at least three sequence motifs that bind NAs, 8 being able to form an interconnected polymer network with NAs. The PrP construct used here, PrP 90-231 , encompasses the second lysine cluster that can interact with NAs, which, when neutralized, results in formation of PrP Sc -like aggregates.…”