title: S-nitrosylation analysis in Brassica juncea apoplast highlights the importance of nitric oxide in cold-stress signaling.
abstract: Reactive nitrogen species (RNS) including nitric oxide (NO) are important components of stress signaling. However, RNS-mediated signaling in the apoplast remains largely unknown. NO production measured in the shoot apoplast of Brassica juncea seedlings showed nonenzymatic nitrite reduction to NO. Thiol pool quantification showed cold-induced increase in the protein (including S-nitrosothiols) as well as non protein thiols. Proteins from the apoplast were resolved as 109 spots on the 2-D gel, while S-nitrosoglutathione-treated (a NO donor), neutravidin-agarose affinity chromatography-purified S-nitrosylated proteins were resolved as 52 spots. Functional categorization after MALDI-TOF/TOF identification showed 41 and 38% targets to be metabolic/cell-wall-modifying and stress-related, respectively, suggesting the potential role(s) of S-nitrosylation in regulating these responses. Additionally, identification of cold-stress-modulated putative S-nitrosylated proteins by nLC-MS/MS showed that only 38.4% targets with increased S-nitrosylation were secreted by classical pathway, while the majority (61.6%) of these were secreted by unknown/nonclassical pathways. Cold-stress-increased dehydroascorbate reductase and glutathione S-transferase activity via S-nitrosylation and promoted ROS detoxification by ascorbate regeneration and hydrogen peroxide detoxification. Taken together, cold-mediated NO production, thiol pool enrichment, and identification of the 48 putative S-nitrosylated proteins, including 25 novel targets, provided the preview of RNS-mediated cold-stress signaling in the apoplast.
authors.affiliation: Molecular Plant Physiology and Proteomics Laboratory, Department of Botany, University of Delhi , Delhi 110007, India.
authors.forename: Ankita
authors.initials: A
authors.lastname: Sehrawat
authors.forename: Renu
authors.initials: R
authors.lastname: Deswal
mesh_terms: D002473:Cell Wall / Q000378:metabolism; D002853:Chromatography, Liquid; D003080:Cold Temperature; D015180:Electrophoresis, Gel, Two-Dimensional; D005110:Extracellular Space / Q000378:metabolism; D008954:Models, Biological; D009149:Mustard Plant / Q000187:drug effects / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism*; D020030:Nitric Oxide Donors / Q000378:metabolism / Q000494:pharmacology; D010940:Plant Proteins / Q000378:metabolism; D020543:Proteome / Q000378:metabolism; D040901:Proteomics / Q000379:methods; D026422:S-Nitrosoglutathione / Q000378:metabolism / Q000494:pharmacology; D026403:S-Nitrosothiols / Q000378:metabolism; D036226:Seedlings / Q000187:drug effects / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013312:Stress, Physiological / Q000502:physiology*; D013438:Sulfhydryl Compounds / Q000378:metabolism; D053719:Tandem Mass Spectrometry
chemical_list: D020030:Nitric Oxide Donors; D010940:Plant Proteins; D020543:Proteome; D026403:S-Nitrosothiols; D013438:Sulfhydryl Compounds; D009569:Nitric Oxide; D026422:S-Nitrosoglutathione