Redox regulation of a guard cell SNF1-related protein kinase in Brassica napus, an oilseed crop

Mengmeng Zhu, Tong Zhang, Wei Ji, Cecilia Silva-Sanchez, Wen Yuan Song, Sarah M. Assmann, Alice C. Harmon, Sixue Chen

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Abstract

Kinase-mediated phosphorylation is a pivotal regulatory process in stomatal responses to stresses. Through a redox proteomics study, a sucrose non-fermenting 1-related protein kinase (SnRK2.4) was identified to be redox-regulated in Brassica napus guard cells upon abscisic acid treatment. There are six genes encoding SnRK2.4 paralogs in B. napus. Here, we show that recombinant BnSnRK2.4-1C exhibited autophosphorylation activity and preferentially phosphorylated the N-terminal region of B. napus slow anion channel (BnSLAC1-NT) over generic substrates. The in vitro activity of BnSnRK2.4-1C requires the presence of manganese (Mn2+). Phosphorylation sites of autophosphorylated BnSnRK2.4-1C were mapped, including serine and threonine residues in the activation loop. In vitro BnSnRK2.4-1C autophosphorylation activity was inhibited by oxidants such as H2O2 and recovered by active thioredoxin isoforms, indicating redox regulation of BnSnRK2.4-1C. Thiol-specific isotope tagging followed by mass spectrometry analysis revealed specific cysteine residues responsive to oxidant treatments. The in vivo activity of BnSnRK2.4-1C is inhibited by 15 min of H2O2 treatment. Taken together, these data indicate that BnSnRK2.4-1C, an SnRK preferentially expressed in guard cells, is redox-regulated with potential roles in guard cell signal transduction.

Original languageEnglish (US)
Pages (from-to)2585-2599
Number of pages15
JournalBiochemical Journal
Volume474
Issue number15
DOIs
StatePublished - Aug 1 2017

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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    Zhu, M., Zhang, T., Ji, W., Silva-Sanchez, C., Song, W. Y., Assmann, S. M., Harmon, A. C., & Chen, S. (2017). Redox regulation of a guard cell SNF1-related protein kinase in Brassica napus, an oilseed crop. Biochemical Journal, 474(15), 2585-2599. https://doi.org/10.1042/BCJ20170070