Anomalous Irreversible “Conversion” of Tag-BFP Fluorescence Spectra in the Presence of Tag-RFP: Possible Model of Prion Interactions

Abstract

Prions are misfolded protein molecules that can propagate by transmitting a misfolded protein state. If the infectious isoform of prion proteins (PrP), known as PrP scrapie (PrPSc), contacts the properly folded PrP called cellular PrP (PrPC), it can stimulate the refolding of normal PrPC into abnormal PrPSc. In the process, the abnormal PrPSc form acts as a template to guide the misfolding of normal PrPC to convert it into more abnormal protein. We encountered similar conversion of fluorescent proteins (FPs) after introducing different FP transgene constructs into several genomic target locations. We used monomeric FPs from Evrogen: TagBFP and RFPs (TagRFP and FusionRed); and Clontech's mCherry.In a variety of targets, across multiple cell lines generated, we noted TagBFP emitting light at RFP wavelengths under certain circumstances. This “conversion” phenomenon existed when the cell line contained both TagBFP and TagRFP expressed at high levels or when they were co-localized. We RFP-tagged α-tubulin (TUBA1B), a highly expressed cytoskeletal protein that polymerizes to form microtubules, using three RFP versions (TagRFP, FusionRed, and mCherry) in cells expressing a TagBFP Segmentation Marker driven off the highly expressed β-actin locus. The complex formed from TagBFP and TagRFP showed an increase in the red channel. In the case when only TagBFP-β-actin fusion was expressed in the absence of any other FP tags, a signal was also present in the red channel. We hypothesize that transient TagBFP-TagRFP heteromerization induces a red-emitting TagBFP conformation. The TagRFP mutations (94% a.a. homology TagBFP) make TagBFP emit in blue. Our results suggest that TagRFP may act as a template to guide the re-folding of TagBFP into a red-emitting conformation. It makes TagBFP-TagRFP pair a convenient tool to model protein-protein interactions during the prion conversion.