Overexpression of a directed mutant of 14-3-3u in Arabidopsis leaves affects phosphorylation and protein content of nitrate reductase

Man Ho Oh, Joan L. Huber, Wei Shen, Gurdeep S. Athwal, Xia Wu, Steven C. Huber

Research output: Contribution to journalArticlepeer-review


The 14-3-3 family of proteins are highly conserved signaling proteins in eukaryotes that bind to their client proteins, usually through specific phosphorylated target sequences. While the 14-3-3 proteins are thought to interact with a wide array of cellular proteins, there have been few studies addressing the in-vivo role of 14-3-3. As one approach to study this in-vivo role, we generated transgenic Arabidopsis plants constitutively overexpressing a directed mutant of 14-3-3 isoform u that inhibits phosphorylated nitrate reductase (pNR) in a largely divalent-cation-independent manner in vitro. The transgenic plants had increased relative phosphorylation of NR at the regulatory Ser-534 site and decreased NR activity measured in the presence of 5 mmol L-1 MgCl2relative to nontransgenic plants. In addition, total NR protein was increased and the protein half-life was increased about two-fold. Two-dimensional difference gel electrophoresis analysis of proteins extracted from leaves of plants expressing the mutant 14-3-3 identified numerous cellular proteins that were altered in abundance. In particular, several b-glucosidase and glutathione S-transferase isoforms were decreased in abundance relative to wild type plants suggesting a possible alteration in stress or defense responses.

Original languageEnglish (US)
Pages (from-to)691-701
Number of pages11
Issue number7
StatePublished - Jul 2009


  • 14-3-3 Protein
  • Arabidopsis thaliana
  • B-glucosidase
  • Glutathione S-transferase
  • Site-directed mutagenesis
  • Two-dimensional difference gel electrophoresis

ASJC Scopus subject areas

  • Ecology, Evolution, Behavior and Systematics
  • Ecology
  • Plant Science


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