Citrus is a globally important, perennial fruit crop whose rhizosphere microbiome is thought to play an important role in promoting citrus growth and health. Here, we report a comprehensive analysis of the structural and functional composition of the citrus rhizosphere microbiome. We use both amplicon and deep shotgun metagenomic sequencing of bulk soil and rhizosphere samples collected across distinct biogeographical regions from six continents. Predominant taxa include Proteobacteria, Actinobacteria, Acidobacteria and Bacteroidetes. The core citrus rhizosphere microbiome comprises Pseudomonas, Agrobacterium, Cupriavidus, Bradyrhizobium, Rhizobium, Mesorhizobium, Burkholderia, Cellvibrio, Sphingomonas, Variovorax and Paraburkholderia, some of which are potential plant beneficial microbes. We also identify over-represented microbial functional traits mediating plant-microbe and microbe-microbe interactions, nutrition acquisition and plant growth promotion in citrus rhizosphere. The results provide valuable information to guide microbial isolation and culturing and, potentially, to harness the power of the microbiome to improve plant production and health.

As an important second messenger, calcium is involved in plant cold stress response, including chilling (<20°C) and freezing (<0°C). In this study, exogenous application of calcium chloride (CaCl 2 ) improved both chilling and freezing stress tolerances, while ethylene glycol-bis-(baminoethyl) ether-N,N,N,N-tetraacetic acid (EGTA) reversed CaCl 2 effects in bermudagrass (Cynodon dactylon (L.) Pers.). Pysiological analyses showed that CaCl 2 treatment alleviated the reactive oxygen species (ROS) burst and cell damage triggered by chilling stress, via activating antioxidant enzymes, non-enzymatic glutathione antioxidant pool, while EGTA treatment had the opposite effects. Additionally, comparative proteomic analysis identified 51 differentially expressed proteins that were enriched in redox, tricarboxylicacid cycle, glycolysis, photosynthesis, oxidative pentose phosphate pathway, and amino acid metabolisms. Consistently, 42 metabolites including amino acids, organic acids, sugars, and sugar alcohols were regulated by CaCl 2 treatment under control and cold stress conditions, further confirming the common modulation of CaCl 2 treatment in carbon metabolites and amino acid metabolism. Taken together, this study reported first evidence of the essential and protective roles of endogenous and exogenous calcium in bermudagrass response to cold stress, partially via activation of the antioxidants and modulation of several differentially expressed proteins and metabolic homeostasis in the process of cold acclimation.Keywords: Antioxidant; bermudagrass; calcium; chilling; freezing; metabolite; proteomic Citation: Shi H, Ye T, Zhong B, Liu X, Chan Z (2014) Comparative proteomic and metabolomic analyses reveal mechanisms of improved cold stress tolerance in bermudagrass (Cynodon dactylon (L.) Pers.) by exogenous calcium.

SummarySeveral eukaryotic Heme-associated proteins (HAPs) have been reported to bind specifically to DNA fragments containing CCAAT-box; however, the physiological functions and direct targets of these HAP proteins in plants remain unclear.In this study, we showed that AtHAP5A as a transcription factor interacted with CCAAT motif in vivo, and AtXTH21, one direct target of AtHAP5A, was involved in freezing stress resistance. The AtHAP5A overexpressing plants were more tolerant, whereas the loss-offunction mutant of AtHAP5A was more sensitive to freezing stress than wild-type plants. Chromatin immunoprecipitation (ChIP) assay demonstrated that AtHAP5A could bind to five fragments that contained CCAAT motifs in the AtXTH21 promoter.Similarly, the AtXTH21 overexpressing plants exhibited improved freezing resistance, while xth21 knockdown mutants displayed decreased freezing resistance. Notably, the modulated freezing resistance of AtHAP5A overexpressing plants and knockout mutant could be reversed by the xth21 mutant and AtXTH21 overexpressing plants, respectively, indicating that AtHAP5A might act upstream of AtXTH21 in freezing stress. Additionally, modulation of AtHAP5A and AtXTH21 expression had the same effects on abscisic acid (ABA) sensitivity and reactive oxygen species (ROS) metabolism.Taken together, these results demonstrated that AtHAP5A modulates freezing stress resistance in Arabidopsis through binding to the CCAAT motif of AtXTH21.