Physiological and Transcriptional Responses of Apocynum venetum to Salt Stress at the Seed Germination Stage.

Apocynum venetum is a semi-shrubby perennial herb that not only prevents saline-alkaline land degradation but also produces leaves for medicinal uses. Although physiological changes during the seed germination of A. venetum in response to salt stress have been studied, the adaptive mechanism to salt conditions is still limited. Here, the physiological and transcriptional changes during seed germination under different NaCl treatments (0-300 mmol/L) were examined. The results showed that the seed germination rate was promoted at low NaCl concentrations (0-50 mmol/L) and inhibited with increased concentrations (100-300 mmol/L); the activity of antioxidant enzymes exhibited a significant increase from 0 (CK) to 150 mmol/L NaCl and a significant decrease from 150 to 300 mmol/L; and the content of osmolytes exhibited a significant increase with increased concentrations, while the protein content peaked at 100 mmol/L NaCl and then significantly decreased. A total of 1967 differentially expressed genes (DEGs) were generated during seed germination at 300 mmol/L NaCl versus (vs.) CK, with 1487 characterized genes (1293 up-regulated, UR; 194 down-regulated, DR) classified into 11 categories, including salt stress (29), stress response (146), primary metabolism (287), cell morphogenesis (156), transcription factor (TFs, 62), bio-signaling (173), transport (144), photosynthesis and energy (125), secondary metabolism (58), polynucleotide metabolism (21), and translation (286). The relative expression levels (RELs) of selected genes directly involved in salt stress and seed germination were observed to be consistent with the changes in antioxidant enzyme activities and osmolyte contents. These findings will provide useful references to improve seed germination and reveal the adaptive mechanism of A. venetum to saline-alkaline soils.

The synergistic effects of sodium and potassium on the xerophyte Apocynum venetum in response to drought stress.

Apocynum venetum is an eco-economic plant species with high adaptability to saline and arid environments. Our previous work has found that A. venetum could absorb large amount of Na+ and maintain high K+ level under saline conditions. To investigate whether K+ and Na+ could simultaneously enhance drought resistance in A. venetum, seedlings were exposed to osmotic stress (-0.2 MPa) in the presence or absence of additional 25 mM NaCl under low (0.01 mM) and normal (2.5 mM) K+ supplying conditions, respectively. The results showed that A. venetum should be considered as a typical K+-efficient species since its growth was unimpaired and possessed a strong K+ uptake and prominent K+ utilization efficiency under K+ deficiency condition. Leaf K+ concentration remained stable or was even significantly increased under osmotic stress in the presence or absence of NaCl, compared with that under control condition, regardless of whether the K+ supply was sufficient or not, and the contribution of K+ to leaf osmotic potential consistently exceeded 37%, indicating K+ is the uppermost contributor to osmotic adjustment of A. venetum. Under osmotic stress, the addition of 25 mM NaCl significantly increase Na+ accumulation in leaves and the contribution of Na+ to osmotic adjustment, thus improving the relative water content, concomitantly, promoting the photosynthetic activity resulting in an enhancement of overall plant growth. These findings suggested that, K+ and Na+ simultaneously play crucial roles in the osmotic adjustment and the maintenance of water status and photosynthetic activity, which is beneficial for A. venetum to cope with drought stress.

Variations in Physiology and Multiple Bioactive Constituents under Salt Stress Provide Insight into the Quality Evaluation of Apocyni Veneti Folium.

As one of the major abiotic stresses, salinity stress may affect the physiology and biochemical components of Apocynum venetum L. To systematically evaluate the quality of Apocyni Veneti Folium (AVF) from the perspective of physiological and the wide variety of bioactive components response to various concentrations of salt stress, this experiment was arranged on the basis of ultra-fast liquid chromatography tandem triple quadrupole mass spectrometry (UFLC-QTRAP-MS/MS) technology and multivariate statistical analysis. Physiological characteristics of photosynthetic pigments, osmotic homeostasis, lipid peroxidation product, and antioxidative enzymes were introduced to investigate the salt tolerance mechanism of AVF under salinity treatments of four concentrations (0, 100, 200, and 300 mM NaCl, respectively). Furthermore, a total of 43 bioactive constituents, including 14 amino acids, nine nucleosides, six organic acids, and 14 flavonoids were quantified in AVF under salt stress. In addition, multivariate statistical analysis, including hierarchical clustering analysis, principal component analysis (PCA), and gray relational analysis (GRA) was employed to systematically cluster, distinguish, and evaluate the samples, respectively. Compared with the control, the results demonstrated that 200 mM and 100 mM salt stress contributed to maintain high quality of photosynthesis, osmotic balance, antioxidant enzyme activity, and the accumulation of metabolites, except for total organic acids, and the quality of AVF obtained by these two groups was better than others; however, under severe stress, the accumulation of the oxidative damage and the reduction of metabolite caused by inefficiently scavenging reactive oxygen species (ROS) lead to lower quality. In summary, the proposed method may provide integrated information for the quality evaluation of AVF and other salt-tolerant Chinese medicines.

[Impact of high salt stress on Apocynum venetum growth and ionic homeostasis].

A pot experiment was conducted in a net room to study the growth responses and related mechanisms of Apocynum venetum treated with different concentrations (100-400 mmol x L(-1)) of NaCl for 30 days. The biomass accumulation, growth rate, root vigor, salt ion content and mineral ion uptake and distribution were measured. Compared with the control, treatment 100 mmol x L(-1) NaCl had lesser effects on the plant dry mass, but decreased the plant fresh mass and growth rate significantly. With increasing NaCl concentration in the medium, the plant dry mass, fresh mass, and growth rate all decreased significantly. The plant root vigor was obviously higher under 100 and 200 mmol x L(-1) NaCl stress, but decreased significantly under 300-400 mmol x L(-1) NaCl stress. With the increase of NaCl concentration in the medium, the Na+ content in A. venetum roots, stems and leaves increased gradually while the K+ content had a slow decrease, the Ca2+ and Mg2+ contents in leaves decreased obviously, and the Ca2+ content in stems and the Mg2+ content in roots increased in different degree. Under NaCl stress, the K+ /Na+, Ca2+/Na+, and Mg2+/Na+ ratios in roots, stems, and leaves decreased markedly, while the selective absorption and transportation of K+ and Ca2+ increased significantly. The stronger ability of salt exclusion and the higher selective absorption and transportation of K+ and Ca2+ were the key adaptive mechanisms of high salt-tolerance of A. venetum.

Molecular cloning and characterization of a salinity stress-induced gene encoding DEAD-box helicase from the halophyte Apocynum venetum.

The genes encoding DEAD-box helicases play a key role in various abiotic stresses, including temperature, light, oxygen, and salt stress. A salt-responsive gene, designated AvDH1, was isolated from the halophyte dogbane (Apocynum venetum) by using suppression subtractive hybridization and RACE (rapid amplification of cDNA ends) PCR. The deduced amino acid sequence has nine conserved helicase motifs of the DEAD-box protein family. The AvDH1 gene is present as a single copy in the dogbane genome. This gene is expressed in response to NaCl and not polyethlene glycol (PEG) nor abscisic acid, and its expression increases with time. The transcription of AvDH1 is also induced by low temperature (4 degrees C), but its accumulation first increases then decreases with time. The purified recombinant protein contains ATP-dependent DNA helicase activity, ATP-independent RNA helicase activity, and DNA- or RNA-dependent ATPase activity. The ATPase activity of AvDH1 is stimulated more by single-stranded DNA than by double-stranded DNA or RNA. These results suggested that AvDH1 belonging to the DEAD-box helicase family is induced by salinity, functions as a typical helicase to unwind DNA and RNA, and may play an important role in salinity tolerance.