Arabidopsis nitrate reductase activity is stimulated by the E3 SUMO ligase AtSIZ1

Abstract

Small ubiquitin-related modifier (SUMO) is a small polypeptide that modulates protein activity and regulates hormone signalling, abiotic and biotic responses in plants. Here we show that AtSIZ regulates nitrogen assimilation in Arabidopsis through its E3 SUMO ligase function. Dwarf plants of siz1-2 flower early, show abnormal seed development and have high salicylic acid content and enhanced resistance to bacterial pathogens. These mutant phenotypes are reverted to wild-type phenotypes by exogenous ammonium but not by nitrate, phosphate or potassium. Decreased nitrate reductase activity in siz1-2 plants resulted in low nitrogen concentrations, low nitric oxide production and high nitrate content in comparison with wild-type plants. The nitrate reductases, NIA1 and NIA2, are sumoylated by AtSIZ1, which dramatically increases their activity. Both sumoylated and non-sumoylated NIA1 and NIA2 can form dimers. Our results indicate that AtSIZ1 positively controls nitrogen assimilation by promoting sumoylation of NRs in Arabidopsis.

Arabidopsis PIAL1 and 2 Promote SUMO Chain Formation as E4-Type SUMO Ligases and Are Involved in Stress Responses and Sulfur Metabolism

“…It is thus plausible that the lack of efficient nitrate reduction is responsible for the accumulation of nitrate. This interpretation is supported by the finding that siz1 and siz1 pial double and triple mutant combinations grow better in presence of additional ammonium ions (Park et al, 2011; data not shown). We conclude from the data that PIAL1 and PIAL2 have only minor influence on nitrate metabolism.…”

“…Measurement of nitrate content further supports this notion ( Figure 6A). Nitrate reductase (NR) is a known substrate of SIZ1-dependent SUMO conjugation and is activated by sumoylation (Park et al, 2011). Increased levels of nitrate in siz1 mutants are consistent with Plants were grown on plates containing 150 mM NaCl as in Figure 2.…”

“…siz1 mutants had increased nitrate content, whereas the content was slightly lowered in pial1 pial2 plants. The increase in siz1 can be related to the finding (Park et al, 2011) that sumoylation of (A) Scheme of constructs and relative activity. In addition to the SP-RING, two potential SIMs were mutated in PIAL2M.…”

“…In the siz1 background, additional mutations in pial1 and pial2 lead to a further, albeit small, reduction in size. All mutant combinations with siz1 are viable (Supplemental Figure 7) and set seed if supplemented with ammonium ions (Park et al, 2011).…”

Overexpression of SIZ1 enhances tolerance to cold and salt stresses and attenuates response to abscisic acid in Arabidopsis thaliana

“…Although there are several SUMO proteins, only two SUMO E3 ligases have been characterized in Arabidopsis-SIZ1 and MMS21/HPY2 (Huang et al 2009;Ishida et al 2009;Miura et al 2005). The siz1 and mms21/hpy2 mutants exhibit pleiotropic phenotypes that involve ABA (abscisic acid), biotic, and Copyright © 2014 The Japanese Society for Plant Cell and Molecular Biology abiotic stress (heat, cold, salt, and drought) responses, nutrient availability, plant development, and flowering (Catala et al 2007;Chen et al 2011;Jin et al 2008;Lee et al 2007;Miura et al 2007b;Miura et al 2011a;Miura and Ohta 2010;Miura et al 2011b;Park et al 2011;Yoo et al 2006). These findings indicate the importance of SUMO E3 ligases in plants.…”