Proteome Analysis Reveals Extensive Light Stress-Response Reprogramming in the Seagrass Zostera muelleri (Alismatales, Zosteraceae) Metabolism

Abstract

Seagrasses are marine ecosystem engineers that are currently declining in abundance at an alarming rate due to both natural and anthropogenic disturbances in ecological niches. Despite reports on the morphological and physiological adaptations of seagrasses to extreme environments, little is known of the molecular mechanisms underlying photo-acclimation, and/or tolerance in these marine plants. This study applies the two-dimensional isoelectric focusing (2D-IEF) proteomics approach to identify photo-acclimation/tolerance proteins in the marine seagrass Zostera muelleri. For this, Z. muelleri was exposed for 10 days in laboratory mesocosms to saturating (control, 200 μmol photons m−2 s−1), super-saturating (SSL, 600 μmol photons m−2 s−1), and limited light (LL, 20 μmol photons m−2 s−1) irradiance conditions. Using LC-MS/MS analysis, 93 and 40 protein spots were differentially regulated under SSL and LL conditions, respectively, when compared to the control. In contrast to the LL condition, Z. muelleri robustly tolerated super-saturation light than control conditions, evidenced by their higher relative maximum electron transport rate and minimum saturating irradiance values. Proteomic analyses revealed up-regulation and/or appearances of proteins belonging to the Calvin-Benson and Krebs cycle, glycolysis, the glycine cleavage system of photorespiration, and the antioxidant system. These proteins, together with those from the inter-connected glutamate-proline-GABA pathway, shaped Z. muelleri photosynthesis and growth under SSL conditions. In contrast, the LL condition negatively impacted the metabolic activities of Z. muelleri by down-regulating key metabolic enzymes for photosynthesis and the metabolism of carbohydrates and amino acids, which is consistent with the observation with lower photosynthetic performance under LL condition. This study provides novel insights into the underlying molecular photo-acclimation mechanisms in Z. muelleri, in addition to identifying protein-based biomarkers that could be used as early indicators to detect acute/chronic light stress in seagrasses to monitor seagrass health.

Additive response to multiple environmental stressors in the seagrass Zostera marina L.

“…LL plants had the lowest respirations rate (per unit biomass and per plant), suggesting that these plants reduced their metabolic rates to save energy. Proteonome analyses have demonstrated that prolonged exposure to shade induces a significant down‐regulation of key enzymes related to photosynthesis, carbohydrate metabolism, and amino acid synthesis in Zostera muelleri (Kumar et al ). Metabolomic studies on Z. marina support these findings: the quantities of sucrose, fructose, and most amino acids declined in plants exposed to shade for ca.…”

“…The response of seagrasses to multiple stressors has lately been addressed experimentally in a number of two-factor studies where potential interactions between a variety of factors have been addressed (Hootsmans et al 1987;Kahn and Durako 2006;Koch et al 2007;de los Santos et al 2010;Collier et al 2011;H€ offle et al 2011;Holmer et al 2011;La Nafie et al 2012;Jiang et al 2013;Villaz� an et al 2013;York et al 2013;Salo and Pedersen 2014;Castorani et al 2015;Villaz� an et al 2015;Hasler-Sheetal et al 2016;Moreno-Mar� ın et al 2016). The conclusions in many of these articles infer a prevalence of synergistic effects in seagrasses when exposed to multiple stressors, although this has rarely been tested explicitly.…”

Development of an Efficient Protein Extraction Method Compatible with LC-MS/MS for Proteome Mapping in Two Australian Seagrasses Zostera muelleri and Posidonia australis

“…Protein (300 μg) was separated by isoelectric focusing (IEF) using a cup-loading method according to Kumar et al ( 2017 ). Immobilized pH gradient (IPG) strips (Bio-Rad, pH 3–10 or 5–8, 11 cm) were passively rehydrated in UTC7 rehydration solution for a minimum of 6 h at room temperature.…”

“…Randomly selected protein spots were excised from gels, trypsin digested, and analyzed by LC/MS/MS according to Kumar et al ( 2017 ). Using an autosampler, connected to a nanoLC system (Tempo Eksigent, USA), 10 μL of the sample was loaded at 20 μL/min with MS loading solvent (2% Acetonitrile + 0.2% Trifluoroacetic Acid) onto a C8 trap column (CapTrap.…”

“…To understand the change in state of the proteins, it is now common to perform a differential display of the proteome under contrasting conditions. Proteomics using advanced mass-spectrometry based approaches have had an increasing impact on the study of terrestrial plant responses to (a) biotic stresses (see references in Kumar et al, 2017 ). However, proteomics in seagrasses is still in its incipient stage for two primary reasons—(1) the lack of efficient protein extraction methods, and (2) limited availability of genomic sequence information.…”