Sato T, Maekawa S, Yasuda S, Domeki Y, Sueyoshi K, Fujiwara M, Fukao Y, Goto DB, Yamaguchi J

The balance between carbon (C) and nitrogen (N) availability is an important determinant for different phases of plant growth, however, detailed mechanisms regulating the C/N response are not well understood. We previously revealed two related ubiquitin ligase, ATL31 and ATL6, that functions in the C/N response in Arabidopsis thaliana. Here, ...

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we applied FLAG tag affinity purification and MS analysis to identify proteins targeted by ATL31, and thus likely involved in regulating the phase transition checkpoint according to C/N status. This analysis revealed that 14-3-3 proteins associated with ATL31, and one of these, 14-3-3χ, was selected for detailed characterization. The interaction between ATL31 and 14-3-3χ was confirmed by yeast two-hybrid and co-immunoprecipitation analyses. In vitro assays showed that 14-3-3χ can act as a target of ATL31, which catalysed the ubiquitination of the recombinant protein. Degradation of 14-3-3χin vivo was also shown to be correlated with ATL31 activity, and to occur in a proteasome-dependent manner. Furthermore, 14-3-3 protein accumulation was induced by a typical shift to high C/N stress conditions in Arabidopsis seedlings and this regulated response required ATL31 and ATL6. It was also demonstrated that the over-expression of 14-3-3χ leads to hypersensitivity of Arabidopsis seedlings to C/N stress conditions. These results indicated that ATL31 functions to target and ubiquitinate 14-3-3 proteins for degradation via the ubiquitin-proteasome system during the response to cellular C/N status.

Date: Jun. 10, 2011

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