Monitoring thioredoxin redox with a genetically encoded red fluorescent biosensor

Abstract

Thioredoxin (Trx) is one of the two major thiol antioxidants playing essential roles in redox homeostasis and signaling. Despite the importance, there is a lack of method in monitoring Trx redox dynamics in live cells, hindering a better understanding of physiological and pathological roles of the Trx redox system. In this work, we developed the first genetically encoded fluorescent biosensor for Trx redox by engineering a redox relay between the active-site cysteines of human Trx1 and rxRFP1—a redox-sensitive red fluorescent protein. We utilized the resultant biosensor—TrxRFP1—to selectively monitor perturbations of Trx redox in various mammalian cell lines. We subcellularly localized TrxRFP1 to image compartmentalized Trx redox changes. We further combined TrxRFP1 with a green fluorescent Grx1-roGFP2 biosensor to simultaneously monitor Trx and glutathione redox dynamics in live cells in response to chemical and physiologically relevant stimuli.

The thioredoxin (Trx) redox state sensor protein can visualize Trx activities in the light/dark response in chloroplasts

“…Although several redox sensor proteins, such as Oba-Q (18), rxYFP (19), rxRFP (20), and Re-Q (21), have been developed, these sensor proteins also cannot detect the redox change in Trx molecules directly. For this purpose, TrxRFP was recently developed as a genetically encoded sensor by fusing rxRFP and human Trx1 (22). This Trx1 was reduced by human Trx reductase and oxidized by human Trx peroxidases in a similar manner as native Trx1.…”

Illuminating pathogen–host intimacy through optogenetics

“…These drugs are also active against Leishmania [28], further advocating the utility of calcium releasers and sensors (Table 1). Other appealing optogenetic approaches involve engineering host cells or parasites to harbor reactive oxygen species generating proteins (RGPs) [29], as well as the biosensors of lipids [25,30,31], polar metabolites [32][33][34], nitric oxide [35], voltage [36], redox [37,38], and pH [39][40][41], each of them tailored to address specific paradigms and individual needs (Table 1).…”

Development of a Sensitive Escherichia coli Bioreporter Without Antibiotic Markers for Detecting Bioavailable Copper in Water Environments

“…Whole-cell bioreporters have gained worldwide attention because they are sensitive, selective, portable, reasonably priced, and easy to handle. In addition, they have a relatively long life and a short response time (Fan et al, 2017;Jiang et al, 2017). Typically, a whole-cell bioreporter is consisted of a promoteroperator, which acts as the sensing element for heavy metal ions, with one or more reporter genes that encode easily detectable proteins (Jiang et al, 2017).…”