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 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 human Trx1 and rxRFP1¾a redox-sensitive red fluorescent protein. We utilized the resultant biosensor¾TrxRFP1¾to selectively monitor chemically induced perturbation of Trx redox in various mammalian cell lines. We further combined TrxRFP1 with a green fluorescent Grx1-roGFP2 biosensor to simultaneously monitor Trx and glutathione redox dynamics in live cells. Our data support that the Trx redox system and the glutathione redox system can be individually perturbed, suggesting that the two major antioxidant systems are kinetically separated to play distinct biological roles.

Monitoring thioredoxin redox in live cells with a genetically encoded red fluorescent protein biosensor

253rd National Meeting (2017)

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