Abstract
S-nitrosoglutathione reductase (GSNOR) is believed to modulate effects of reactive oxygen and nitrogen species through catabolism of S-nitrosoglutathione (GSNO). We combined bioinformatics of plant GSNOR genes, localization of GSNOR in Arabidopsis thaliana, and microarray analysis of a GSNOR null mutant to gain insights into the function and regulation of this critical enzyme in nitric oxide (NO) homeostasis. GSNOR-encoding genes are known to have high homology across diverse eukaryotic taxa, but contributions of specific conserved residues have not been assessed. With bioinformatics and structural modeling, we show that plant GSNORs likely localize to the cytosol, contain conserved, solvent-accessible cysteines, and tend to be encoded by a single gene. Arabidopsis thaliana homozygous for GSNOR loss-of-function alleles exhibited defects in stem and trichome branching, and complementation with Green fluorescent protein (GFP) -tagged GSNOR under control of the native promoter quantitatively rescued these phenotypes. GSNOR-GFP showed fluorescence throughout Arabidopsis seedlings, consistent with ubiquitous expression of the protein, but with especially high fluorescence in the root tip, apical meristem, and flowers. At the cellular level we observed cytosolic and nuclear fluorescence, with exclusion from the nucleolus. Microarray analysis identified 99 up- and 170 down-regulated genes (≥2-fold; p ≤ 0.01) in a GSNOR null mutant compared to wild type. Six members of the plant specific, ROXY glutaredoxins and three BHLH transcription factors involved in iron homeostasis were strongly upregulated, supporting a role for GSNOR in redox and iron metabolism. One third of downregulated genes are linked to pathogen resistance, providing further basis for the reported pathogen sensitivity of GSNOR null mutants. Together, these findings indicate GSNOR regulates multiple developmental and metabolic programs in plants and offer insight into putative routes of post-translational GSNOR regulation.
Highlights
In plants, biological processes ranging from leaf stomatal closure to auxin perception in roots and pathogen infection involve nitric oxide (NO) (Neill et al, 2002; Floryszak-Wieczorek et al, 2007; Lozano-Juste and Leon, 2011; Terrile et al, 2012)
Because cysteines can serve as key post-translational regulatory sites being modified by nitrosation, glutathionylation, or reversible oxidation, we analyzed the conservation of the nine extra non-zinc-coordinating cysteine residues in Arabidopsis GSNO by S-nitrosoglutathione reductase (GSNOR)
We examined the position of ex-zinc cysteines in the Arabidopsis GSNOR structure (PDB 4JJI, via (PyMOL) and found that three were solvent accessible (Cys-10, Cys-271, and Cys-370, Figures 2A–C), two of which—Cys-10 and Cys-271—are positionally conserved even in the human sequence
Summary
Biological processes ranging from leaf stomatal closure to auxin perception in roots and pathogen infection involve nitric oxide (NO) (Neill et al, 2002; Floryszak-Wieczorek et al, 2007; Lozano-Juste and Leon, 2011; Terrile et al, 2012). While NO itself is ostensibly active, it is thought to be transmitted to distal targets via low molecular weight S-nitrosothiols (SNOs), of which the glutathione (GSH) adduct S-nitrosoglutathione (GSNO) is the most abundant (Broniowska et al, 2013; Corpas et al, 2013). Catabolism of GSNO by S-nitrosoglutathione reductase (GSNOR) is common to eukaryotes and many bacteria and is believed to be responsible for this regulation of GSNO levels (Liu et al, 2001; Staab et al, 2008). While GSNO catabolism has been observed with Cu-Zn superoxide dismutase, GSH peroxidase, xanthine oxidase, and human carbonyl reductase 1 (CR1), the former three enzymes merely regenerate NO [summarized in Broniowska et al (2013)], and residues critical to interaction between CR1 and GSH adducts are not conserved in plants (Bateman et al, 2008). GSNOR is considered the primary catalyst for GSNO catabolism in plants
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
