Assessment of the changes in growth, photosynthetic traits and gene expression in Cynodon dactylon against drought stress.

Drought stress considered a key restrictive factor for a warm-season bermudagrass growth during summers in China. Genotypic variation against drought stress exists among bermudagrass (Cynodon sp.), but the selection of highly drought-tolerant germplasm is important for its growth in limited water regions and for future breeding. Our study aimed to investigate the most tolerant bermudagrass germplasm among thirteen, along latitude and longitudinal gradient under a well-watered and drought stress condition. Current study included high drought-resistant germplasm, "Tianshui" and "Linxiang", and drought-sensitive cultivars; "Zhengzhou" and "Cixian" under drought treatments along longitude and latitudinal gradients, respectively. Under water deficit conditions, the tolerant genotypes showed over-expression of a dehydrin gene cdDHN4, antioxidant genes Cu/ZnSOD and APX which leads to higher antioxidant activities to scavenge the excessive reactive oxygen species and minimizing the membrane damage. It helps in maintenance of cell membrane permeability and osmotic adjustment by producing organic osmolytes. Proline an osmolyte has the ability to keep osmotic water potential and water use efficiency high via stomatal conductance and maintain transpiration rate. It leads to optimum CO2 assimilation rate, high chlorophyll contents for photosynthesis and elongation of leaf mesophyll, palisade and thick spongy cells. Consequently, it results in elongation of leaf length, stolon and internode length; plant height and deep rooting system. The CdDHN4 gene highly expressed in "Tianshui" and "Youxian", Cu/ZnSOD gene in "Tianshui" and "Linxiang" and APX gene in "Shanxian" and "Linxiang". The genotypes "Zhongshan" and "Xiaochang" showed no gene expression under water deficit conditions. Our results indicate that turfgrass show morphological modifications firstly when subjected to drought stress; however the gene expression is directly associated and crucial for drought tolerance in bermudagrass. Hence, current research has provided excellent germplasm of drought tolerant bermudagrass for physiological and molecular study and future breeding.

Leaf Cuticular Waxes of Bermudagrass Response to Environment-Driven Adaptations of Climate Effect Inferred from Latitude and Longitude Gradient in China.

Bermudagrass (Cynodon dactylon) is a widely used warm season lawn grass. Cuticular wax covering the surface of plant leaves plays an important role in helping plants resist biotic and abiotic stresses. We analyzed the changes of cuticle wax in 25 bermudagrass populations from different longitude and latitude gradients, in order to verify how environmental conditions affect the structure and chemical composition of cuticle wax. Five wax components were identified, including alkanes, esters, alkenes, aldehydes and primary alcohols. The wax characteristics were divided into two principal components, explaining 58.2 % and 66.7 % of the total variability in latitude and longitude, even some populations had a certain correlation with each other. Pearson correlation analysis further showed that the total wax coverage, wax component content and antioxidant enzyme activity of bermudagrass populations on the latitudinal gradient had different responses to environmental factors. Finally, nineteen key genes involved in wax biosynthesis, redox and photosynthesis were identified and verified by RT-qPCR. The results showed that the responses of bermudagrass in different populations to climate change were quite different, which was of great significance for the evolution of bermudagrass populations.

A Novel Beta-Glucosidase Gene for Plant Type Was Identified by Genome-Wide Association Study and Gene Co-Expression Analysis in Widespread Bermudagrass.

Bermudagrass (Cynodon spp.) is one of the most widely distributed warm-season grasses globally. The growth habits and plant type of bermudagrass are strongly associated with the applied purpose of the landscape, livestock, and eco-remediation. Therefore, persistent efforts are made to investigate the genetic basis of plant type and growth habits of bermudagrass. Here, we dissect the genetic diversity of 91 wild bermudagrass resources by genome-wide association studies (GWAS) combined with weighted gene co-expression analysis (WGCNA). This work is based on the RNA-seq data and the genome of African bermudagrass (Cynodon transvaalensis Burtt Davy). Sixteen reliable single-nucleotide polymorphisms (SNPs) in transcribed regions were identified to be associated with the plant height and IAA content in diverse bermudagrass by GWAS. The integration of the results from WGCNA indicates that beta-glucosidase 31 (CdBGLU31) is a candidate gene underlying a G/A SNP signal. Furthermore, both qRT-PCR and correlation coefficient analyses indicate that CdBGLU31 might play a comprehensive role in plant height and IAA biosynthesis and signal. In addition, we observe lower plant height in Arabidopsis bglu11 mutants (homologs of CdBGLU31). It uncovers the breeding selection history of different plant types from diverse bermudagrass and provides new insights into the molecular function of CdBGLU31 both in plant types and in IAA biosynthetic pathways.

Preparation of Highly Substituted Sulfated Alfalfa Polysaccharides and Evaluation of Their Biological Activity.

Alfalfa polysaccharides (AP) receive wide attention in the field of medicine, because of their anti-inflammatory property. However, AP has high molecular weight and poor water solubility, resulting in low biological activity. We wanted to obtain highly bioactive alfalfa polysaccharides for further research. Herein, we successfully synthesized highly substituted sulfated alfalfa polysaccharides (SAP) via the chlorosulfonic acid (CSA)-pyridine (Pyr) method, which was optimized using response surface methodology (RSM). Under the best reaction conditions, that is, the reaction temperature, time, and ratio of CSA to Pyr being 55 °C, 2.25 h, and 1.5:1, respectively, the maximum degree of substitution of SAP can reach up to 0.724. Fourier transform infrared spectroscopy also confirmed the existence of sulfonic acid groups on SAP. Despite the increased average molecular weight of SAP, its water solubility is improved, which is beneficial for its biological activity. Further in vitro results showed that SAP exhibited better antioxidant activity and antibacterial ability than AP. Besides, the former can efficiently enhance the viability of oxidatively stressed intestinal epithelial cells compared with the latter. Furthermore, SAP has the potential to inhibit obesity. It is concluded that sulfation modification could improve the antioxidant, antibacterial, bovine intestinal epithelial cells' proliferation-promoting, and the obesity inhibition abilities of AP. The improvement of AP biological activity may provide references for the utilization of plant extracts that have weaker biological activity.

Morphological variation in Cynodon dactylon (L.) Pers., and its relationship with the environment along a longitudinal gradient.

BACKGROUND: Geographical variation in morphological traits may reflect evolutionary patterns of morphological adaptability along environmental gradients. Comprehensive information on longitudinal patterns of morphological trait variation is very meaningful to explore morphological diversity and evolutionary trends in widespread bermudagrass. METHODS: To explore the spatial patterns of morphological traits, we investigated 10 morphological traits of bermudagrass and 10 soil nutrient indexes and collected local climate data for 13 different regions from 119°E to 105°E along the latitude 34°N. RESULTS: Considerable variations in morphological traits were observed at different longitudes, and the variations in most of the evaluated traits within populations were lower than those among populations. All of the 13 different longitudinal sites were divided into three groups based on morphological traits by cluster analysis. The major sources of diversity at the different longitudes were leaf length of the erect shoot, leaf width of the erect shoot, and the internode lengths of the erect shoot and stolon as determined by principal component analysis. Pearson correlation analysis also indicated that longitude was significantly and negatively correlated with these traits as well. Mean average rainfall was significantly correlated with leaf length of the erect shoot and the internode lengths of the erect shoot and stolon, while mean average temperature was only significantly correlated with internode length of the erect shoots. Available sulfur was significantly correlated with internode length of the erect shoot, plant height, and reproductive branch height, while the exchangeable Ca was significantly correlated with internode lengths of the erect shoot and stolon. Soil pH was significantly correlated with the internode length of the stolon. Longitude is an important factor that affects morphological trait variation in wild bermudagrass, and the leaves of the erect shoot and the internode length enlarged significantly with the collection sites moving from east to west. CONCLUSION: Different combinations and interactions of environmental factors (soil and climate) along a longitudinal gradient may have strong effects on one or more morphological traits of bermudagrass.

Unbiased phosphoproteome profiling uncovers novel phosphoproteins and phosphorylation motifs in bermudagrass stolons.

As a widely used turfgrass species, bermudagrass (Cynodon dactylon L.) can be easily propagated through colonial growth of stolons. Previous studies collectively revealed that exotic environmental factors and intrinsic hormones and genes are all involved in the differentiation, development, and diageotropical growth of stolons. However, the detailed molecular mechanism how environmental and hormone signals regulate the gene expression and biochemical activities in bermudagrass stolons remains unclear. In this study, we observed that reversible phosphorylation modification plays important roles in normal growth and physiological functions of bermudagrass stolons. LC-MS/MS analyses of the total protein extracts of bermudagrass stolons without preliminary phosphopeptide-enrichment successfully identified 646 nonredundant phosphorylation sites and 485 phosphoproteins. The phosphoproteins were significantly enriched in protein phosphorylation regulation and starch metabolism processes. Motif-X analyses further revealed that phosphoproteins containing novel phosphorylation motifs might be involved in transcription regulation of bermudagrass stolons. These results greatly expanded our understanding of the growth and development of bermudagrass stolons at the post-translational level.

Variations in morphological traits of bermudagrass and relationship with soil and climate along latitudinal gradients.

BACKGROUND: This complex environmental heterogeneity coupled with the long-standing history offers scenario suitable for and favoring the evolution and existence of variation of morphological traits. METHODS: In this study, we measured 10 morphological traits of 310 Cynodon dactylon individuals sampled at 16 different locations along latitudinal gradients between 22°35'N and 36°18'N to reveal phenotypic plasticity influenced by latitude. In addition, the relationships between morphological variation and soil nutrient and climate factors were analyzed. RESULTS: Analysis of variance, divesity examination and Mantel correlation test detected a significant effect of latitude on morphological traits. Cluster analysis and principal component analysis clearly separated the selected populations into four groups according to latitude. Larger morphological sizes of C. dactylon appeared at the low- and high-latitude regions. Correlation analysis indicated that high morphological variations were significantly correlated with climate factors and soil nutrient. CONCLUSION: This study suggests morphological variation of wild bermudagrass is greatly influenced by latitude as well as soil and climate, which could be useful resources for genetic studies and evolution.

The regulation of alfalfa saponin extract on key genes involved in hepatic cholesterol metabolism in hyperlipidemic rats.

To investigate the cholesterol-lowering effects of alfalfa saponin extract (ASE) and its regulation mechanism on some key genes involved in cholesterol metabolism, 40 healthy 7 weeks old male Sprague Dawley (SD) rats were randomly divided into four groups with 10 rats in each group: control group, hyperlipidemic group, ASE treatment group, ASE prevention group. The body weight gain, relative liver weight and serum lipid 1evels of rats were determined. Total cholesterol (TC) and total bile acids (TBA) levels in liver and feces were also measured. Furthermore, the activity and mRNA expressions of Hmgcr, Acat2, Cyp7a1 and Ldlr were investigated. The results showed the following: (1) The abnormal serum lipid levels in hyperlipidemic rats were ameliorated by ASE administration (both ASE prevention group and treatment group) (P<0.05). (2) Both ASE administration to hyperlipidemic rats significantly reduced liver TC and increased liver TBA level (P<0.05). TC and TBA levels in feces of hyperlipidemic rats were remarkably elevated by both ASE administration (P<0.05). (3) mRNA expressions of Hmgcr and Acat2 in the liver of hyperlipidemic rats were remarkably down-regulated (P<0.05), as well as mRNA expressions of Cyp7a1 and Ldlr were dramatically up-regulated by both ASE administration (P<0.05). The activities of these enzymes also paralleled the observed changes in mRNA levels. (4) There was no significant difference between ASE treatment and ASE prevention group for most parameters evaluated. Our present study indicated that ASE had cholesterol-lowering effects. The possible mechanism could be attributed to (1) the down-regulation of Hmgcr and Acat2, as well as up-regulation of Cyp7a1 and Ldlr in the liver of hyperlipidemic rats, which was involved in cholesterol biosynthesis, uptake, and efflux pathway; (2) the increase in excretion of cholesterol. The findings in our study suggested ASE had great potential usefulness as a natural agent for treating hyperlipidemia.