New insights into physiological effects of anoxia under darkness on the iconic seagrass Zostera marina based on a combined analysis of transcriptomics and metabolomics.

Coastal hypoxia/anoxia is a major emerging threat to global coastal ecosystems. Macroalgae blooms of tens of kilometers are often observed in open waters. These blooms not only cause a lack of oxygen, but also benthic light limitation. We explored the physiological responses of Zostera marina L. to anoxia under darkness. After exposing Z. marina to anoxia under darkness for 72 h, we measured the elongation of leaves and the decrease in maximal quantum yield of photosystem II (Fv/Fm), and investigated the transcriptomic and metabolomic responses to anoxic stress based on RNA-sequencing and liquid chromatography-mass spectrometry (LC-MS) technology. The results showed that anoxic stress significantly reduced the leaf Fv/Fm, and had a significant negative effect on the photosynthesis and growth of Z. marina. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis of up-regulated differentially expressed genes (DEGs) showed that glycolysis was the most significant enrichment pathway (p < 0.001), and most of the important products in glycolysis were significantly up-regulated. This indicated that the glycolysis process of anaerobic respiration is promoted under anoxia. The metabolite results also showed that glyceraldehyde 3-phosphate in the glycolysis pathway was significantly up-regulated. Moreover, three genes encoding sucrose synthase (gene-ZOSMA_310G00150, gene-ZOSMA_81G00980, and gene-ZOSMA_8G00730) and one gene encoding alpha-amylase (gene-ZOSMA_95G00270) were significantly up-regulated, providing the sugar basis for the subsequent increase in glycolysis. Furthermore, gene-encoding oxoglutarate dehydrogenase, the rate-limiting step of the tricarboxylic acid (TCA) cycle, was significantly down-regulated, indicating that this cycle was inhibited under anoxia. Metabolomic results showed that L-tryptophan, L-phenylalanine, and DL-leucine were significantly up-regulated. Only significantly decreased glutamate and non-significantly decreased glutamine, substances consumed in alanine and γ-aminobutyric acid (GABA) shunt mechanisms, were detected in the leaves, while GABA and alanine were not detected. The results of this study show that anoxic stress induces a programmed transcriptomic and metabolomic response in seagrass, most likely reflecting a complex strategy of acclimation and adaptation in seagrass to resist anoxic stress.