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Multiple cis-regulatory elements and the yeast sulphur regulatory network are required for the regulation of the yeast glutathione transporter, Hgt1p

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Abstract

HGT1 encodes a high-affinity glutathione transporter in the yeast Saccharomyces cerevisiae that is induced under sulphur limitation. The present work demonstrates that repression by organic sulphur sources is under the control of the classic sulphur regulatory network, as seen by the absence of expression in a met4Δ background. Cysteine appeared to be the principal regulatory molecule, since elevated levels were seen in str4Δ strains (deficient in cysteine biosynthesis) that could be repressed by elevated levels of cysteine, but not by methionine or glutathione. Investigations into cis-regulatory elements revealed that the previously described motif, a 9-bp cis element, CCGCCACAC, located at the −356 to −364 region of the promoter could in fact be refined to a 7-bp CGCCACA motif that is also repeated at −333 to −340. The second copy of this motif was essential for activity, since mutations in the core region of the second copy completely abolished activity and regulation by sulphur sources. Activity, but not regulation, could be restored by reintroducing an additional copy upstream of the first copy. A third region, GCCGTCTGCAAGGCA, conserved in the HGT1 promoters of the different Saccharomyces spp, was observed at −300 to −285 but, while mutations in this region did not lead to any loss in repression, the basal and induced levels were significantly increased. In contrast to a previous report, no evidence was found for regulation by the VDE endonuclease. The strong repression at the transport level by glutathione seen in strains overexpressing HGT1 was due to a glutathione-dependent toxicity in these cells.

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Acknowledgements

We thank Raj Kumar for technical assistance and Jaspreet Kaur for help in carrying out the cadmium and diamide experiments. We thank Dr. Tom Stevens (University of Oregon, Eugene, Ore.), Dr. Johanssen (Carlsberg Research Laboratory, Copenhagen Valby, Denmark) and Dr. Fred Gimble (University of Texas, Austin, Tex.) for the gift of strains and plasmids. This work was supported by Grant-in-Aid projects to A.K.B. from the Department of Biotechnology, Government of India, and a Grant-in-Aid project from the Indo-French Centre for the Promotion of Advanced Research (2700-B1) to A.K.B. and S.D. C.V.S. was the recipient of a Research Fellowship from the Council of Scientific and Industrial Research, Government of India.

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Authors and Affiliations

  1. Institute of Microbial Technology, Sector 39-A, Chandigarh, 160 036, India

    Chittur V. Srikanth, Purva Vats & Anand K. Bachhawat

  2. UMR CNRS 6161, Transport des Assimilats, Laboratoire de Physiologie, Biochimie et Biologie Moléculaires Végétales, Bâtiment Botanique, UFR Sciences, University of Poitiers, 40 Avenue du Recteur Pineau, Poitiers, 86022 Cedex, France

    Andrée Bourbouloux & Serge Delrot

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  1. Chittur V. Srikanth
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  2. Purva Vats
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  3. Andrée Bourbouloux
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  4. Serge Delrot
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  5. Anand K. Bachhawat
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Correspondence to Anand K. Bachhawat.

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Communicated by S. Hohmann

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Srikanth, C.V., Vats, P., Bourbouloux, A. et al. Multiple cis-regulatory elements and the yeast sulphur regulatory network are required for the regulation of the yeast glutathione transporter, Hgt1p. Curr Genet 47, 345–358 (2005). https://doi.org/10.1007/s00294-005-0571-7

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