Abstract
Reactive oxygen species trigger cellular responses by activation of stress-responsive mitogen-activated protein kinase (MAPK) signalling pathways1,2. Reversal of MAPK activation requires the transcriptional induction of specialized cysteine-based phosphatases that mediate MAPK dephosphorylation3. Paradoxically, oxidative stresses generally inactivate cysteine-based phosphatases by thiol modification and thus could lead to sustained or uncontrolled MAPK activation4,5. Here we describe how the stress-inducible MAPK phosphatase, Sdp1, presents an unusual solution to this apparent paradox by acquiring enhanced catalytic activity under oxidative conditions. Structural and biochemical evidence reveals that Sdp1 employs an intramolecular disulphide bridge and an invariant histidine side chain to selectively recognize a tyrosine-phosphorylated MAPK substrate. Optimal activity critically requires the disulphide bridge, and thus, to the best of our knowledge, Sdp1 is the first example of a cysteine-dependent phosphatase that couples oxidative stress with substrate recognition. We show that Sdp1, and its paralogue Msg5, have similar properties and belong to a new group of phosphatases unique to yeast and fungal taxa.
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Acknowledgements
G.C.F. was supported by a studentship from GlaxoSmithKline. N.Q.M. and S.M.K. are funded by CR-UK. M.J.R.S. is supported by BBSRC. We thank M. Toledano for experimental advice, T. Kuno for the Pmp1 plasmid and M. Way for critical comments on the manuscript. We also thank M. Cobb for the gift of the ERKpTpY expression vector, D. Thiele and J.-S. Hahn for yeast strains and Sdp1 expression plasmids, and A. Stewart, S. Tilley and J. Denu for assistance with early experiments.
Author Contributions G.C.F. and N.Q.M. determined the apo and sulphate-bound structures. M.S. determined the phosphotyrosine structure and carried out pNPP and ERK2 assays and blots. D.C.B. and N.Q.M. refined the structures. P.P.K. measured the free -SH content and reproduced all kinetic experiments. S.M.K. and M.J.R.S. designed the in vivo assays. M.C, M.J.D, V.M., and R.J.D. carried out in vivo assays using Myc-tagged Sdp1 proteins. S.H. and N.T carried out the mass spectrometry. S.M.K. and N.Q.M planned the project and designed the experiments. N.Q.M., S.M.K. and D.C.B. wrote the paper.
Coordinates have been deposited at the PDB with accession codes 2j16 (apo/sulphate-bound Sdp1) and 2j17 (phosphotyrosine–Sdp1 complex)
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Fox, G., Shafiq, M., Briggs, D. et al. Redox-mediated substrate recognition by Sdp1 defines a new group of tyrosine phosphatases. Nature 447, 487–492 (2007). https://doi.org/10.1038/nature05804
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DOI: https://doi.org/10.1038/nature05804
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