The Chlorella vulgaris S-Nitrosoproteome under Nitrogen-Replete and -Deplete Conditions

Citation: Henard CA, Guarnieri MT and Knoshaug EP (2017) The Chlorella vulgaris S-Nitrosoproteome under Nitrogen-Replete and-Deplete Conditions. Oleaginous microalgae synthesize and accumulate large quantities of lipids that are promising feedstocks for the production of biofuels (Hu et al. The algal species Chlorella vulgaris accumulates triacylglycerides that dominate its cellular composition (>60% lipid based on dry cell weight) when cultured in medium lacking a nitrogen source (Guarnieri et al., 2011; Ikaran et al., 2015), which is a " lipid trigger " in an array of microalgae. As such, C. vulgaris represents a model algal species for examination of lipid accumulation mechanisms and a potential deployment organism in industrial algal biofuels applications. C. vulgaris has been extensively characterized biochemically and physiologically (Converti et al., 2009; Liang et al., 2009), and de novo-generated transcriptomic and proteomic datasets have indicated that post-transcriptional and-translational mechanisms likely govern lipid accumulation in response to nitrogen starvation (Guarnieri et al., 2011, 2013). However, the specific mechanisms underlying lipid biosynthesis in response to nitrogen stress remain elusive. Nitric oxide (NO) has received much attention as a signaling molecule due to its ability to react specifically with a limited number of biomolecules, primarily mediating physiological changes by modifying proteins in diverse domains of life, including plants, animals, and prokaryotes. This dia-tomic radical targets [Fe–S] clusters of dehydratases in several central metabolic pathways and can react with redox active sulfhydryls in cysteines to produce S-nitrosothiols. Protein S-nitrosylation by NO has been shown to play important roles in an array of cellular responses in organisms ranging from prokaryotes to metazoans. Previous studies have identified several proteins as targets of NO in higher plants, confirming S-nitrosylation as a key post-translational mechanism governing cell signaling in these organisms (Zaffagnini et al., 2016). Additional studies in the green microalga Chlamydomonas reinhardtii have identified hundreds of S-nitrosylated proteins after treatment with S-nitrosoglutathione (Samuel et al., 2014), supporting S-nitrosylation as a widespread post-translational mechanism employed by autotrophic organisms. Based on the nitrate reductase-dependent NO production observed in higher plants and micro-algae (Sakihama et al., 2002; Mur et al., 2013), we evaluated whether NO is produced by C. vulgaris when cultured in medium with nitrate as the sole nitrogen source. Using the NO-reactive fluoro-phore 4,5-diaminofluorescein, we detected NO produced by C vulgaris during logarithmic growth in modified Bold's Basal Medium (BBM) supplemented with 3 mM sodium nitrate (Figure 1). The NO detected in algae exposed to nitrite in …