TY - JOUR

T1 - Sulfurtransferase and thioredoxin specifically interact as demonstrated by bimolecular fluorescence complementation analysis and biochemical tests

AU - Henne, Melina

AU - König, Nicolas

AU - Triulzi, Tiziana

AU - Baroni, Sara

AU - Forlani, Fabio

AU - Scheibe, Renate

AU - Papenbrock, Jutta

N1 - Funding information: We would like to thank Pamela von Trzebiatowski and Silke Walter for their excellent technical assistance. Anshu Kuriakose, Vellore, India, produced some of the mutant thioredoxin clones as a part of her MSc thesis. We acknowledge the gift of recombinant thioredoxin proteins for initial biochemical analysis from Yves Meyer, Perpignan, France, and the gift of split-YFP vectors from Klaus Harter, Tübingen, Germany. The project was originally started in cooperation with Simone Holtgrefe, Osnabrück, and Divya Bagchi, Jabalpur, India. Christian Boestfleisch, Hannover, helped to improve Fig. 2 . The work was supported financially by grants from the Deutsche Forschungsgemeinschaft to J.P. ( PA 764/1-4 and 1-5), by the Università degli Studi di Milano to T.T., and by the DAAD (Vigoni 0815171 ).

PY - 2015/12/1

Y1 - 2015/12/1

N2 - Sulfurtransferases (Strs) and thioredoxins (Trxs) are members of large protein families. Trxs are disulfide reductases and play an important role in redox-related cellular processes. They interact with a broad range of proteins. Strs catalyze the transfer of a sulfur atom from a suitable sulfur donor to nucleophilic sulfur acceptors in vitro, but the physiological roles of these enzymes are not well defined. Several studies in different organisms demonstrate protein-protein interactions of Strs with members of the Trx family. We are interested in investigating the specificity of the interaction between Str and Trx isoforms. In order to use the bimolecular fluorescence complementation (BiFC), several Str and Trx sequences from Arabidopsis thaliana were cloned into the pUC-SPYNE and pUC-SPYCE split-YFP vectors, respectively. Each couple of plasmids containing the sequences for the putative interaction partners were transformed into Arabidopsis protoplasts and screened using a confocal laser scanning microscope. Compartment- and partner-specific interactions could be observed in transformed protoplasts. Replacement of cysteine residues in the redox-active site of Trxs abolished the interaction signal. Therefore, the redox site is not only involved in the redox reaction but also responsible for the interaction with partner proteins. Biochemical assays support a specific interaction among Strs and certain Trxs. Based on the results obtained, the interaction of Strs and Trxs indicates a role of Strs in the maintenance of the cellular redox homeostasis.

AB - Sulfurtransferases (Strs) and thioredoxins (Trxs) are members of large protein families. Trxs are disulfide reductases and play an important role in redox-related cellular processes. They interact with a broad range of proteins. Strs catalyze the transfer of a sulfur atom from a suitable sulfur donor to nucleophilic sulfur acceptors in vitro, but the physiological roles of these enzymes are not well defined. Several studies in different organisms demonstrate protein-protein interactions of Strs with members of the Trx family. We are interested in investigating the specificity of the interaction between Str and Trx isoforms. In order to use the bimolecular fluorescence complementation (BiFC), several Str and Trx sequences from Arabidopsis thaliana were cloned into the pUC-SPYNE and pUC-SPYCE split-YFP vectors, respectively. Each couple of plasmids containing the sequences for the putative interaction partners were transformed into Arabidopsis protoplasts and screened using a confocal laser scanning microscope. Compartment- and partner-specific interactions could be observed in transformed protoplasts. Replacement of cysteine residues in the redox-active site of Trxs abolished the interaction signal. Therefore, the redox site is not only involved in the redox reaction but also responsible for the interaction with partner proteins. Biochemical assays support a specific interaction among Strs and certain Trxs. Based on the results obtained, the interaction of Strs and Trxs indicates a role of Strs in the maintenance of the cellular redox homeostasis.

KW - Arabidopsis thaliana

KW - Bimolecular fluorescence complementation

KW - Protein interaction

KW - Thioredoxin

UR - http://www.scopus.com/inward/record.url?scp=84945192378&partnerID=8YFLogxK

U2 - 10.1016/j.fob.2015.10.001

DO - 10.1016/j.fob.2015.10.001

M3 - Article

AN - SCOPUS:84945192378

VL - 5

SP - 832

EP - 843

JO - FEBS OPEN BIO

JF - FEBS OPEN BIO

SN - 2211-5463

ER -