Prion protein transcription is auto-regulated through dynamic interactions with G-quadruplex motifs in its own promoter.

Cellular prion protein (PrP) misfolds into an aberrant and infectious scrapie form (PrPSc) that lead to fatal transmissible spongiform encephalopathies (TSEs). Association of prions with G-quadruplex (GQ) forming nucleic acid motifs has been reported, but implications of these interactions remain elusive. Herein, we show that the promoter region of the human prion gene (PRNP) contains two putative GQ motifs (Q1 and Q2) that assume stable, hybrid, intra-molecular quadruplex structures and bind with high affinity to PrP. Here, we investigate the ability of PrP to bind to the quadruplexes in its own promoter. We used a battery of techniques including SPR, NMR, CD, MD simulations and cell culture-based reporter assays. Our results show that PrP auto-regulates its expression by binding and resolving the GQs present in its own promoter. Furthermore, we map this resolvase-like activity to the N-terminal region (residues 23-89) of PrP. Our findings highlight a positive transcriptional-translational feedback regulation of the PRNP gene by PrP through dynamic unwinding of GQs in its promoter. Taken together, our results shed light on a yet unknown mechanism of regulation of the PRNP gene. This work provides the necessary framework for a plethora of studies on understanding the regulation of PrP levels and its implications in prion pathogenesis.

Heterologous expression of an engineered protein domain acts as chaperone and enhances thermotolerance of Escherichia coli.

Heat shock proteins (HSPs) are known to confer protection to the stressed cells by rescuing vital host cell proteins. In the present study we have demonstrated that heterologous expression of N-terminal domain of hyperthermophilic L-asparaginase (NPfA) confers thermotolerance to E. coli. The recombinant expression of NPfA enabled E. coli to demonstrate typical growth behavior at 52°C and survive a thermal shock up to 62°C, both being the highest reported temperatures for growth and heat shock survival. To understand the basis of protection proteome analysis of these cells was carried out which showed that NPfA guards a battery of proteins, especially related to gene regulations and repair, providing definite survival advantage to the stressed cells. Thus NPfA a non-canonical, non-natural chaperone has been shown to render E. coli cells with selective growth advantage under extremes of conditions. We propose that such modified, heat stabilized hosts could be utilized in developing heat-induced expression systems as well for the recombinant expression of thermophilic proteins.