Overexpression of MsNIP2 improves salinity tolerance in Medicago sativa.

Alfalfa (Medicago sativa) is one of the most widely cultivated forage crops in the world. However, alfalfa yield and quality are adversely affected by salinity stress. Nodulin 26-like intrinsic proteins (NIPs) play essential roles in water and small molecules transport and response to salt stress. Here, we isolated a salt stress responsive MsNIP2 gene and demonstrated its functions by overexpression in alfalfa. The open reading frame of MsNIP2 is 816 bp in length, and it encodes 272 amino acids. It has six transmembrane domains and two NPA motifs. MsNIP2 showed high identity to other known NIP proteins, and its tertiary model was similar to the crystal structure of OsNIP2-1 (7cjs) tetramer. Subcellular localization analysis showed that MsNIP2 protein fused with green fluorescent protein (GFP) was localized to the plasma membrane. Transgenic alfalfa lines overexpressing MsNIP2 showed significantly higher height and branch number compared with the non-transgenic control. The POD and CAT activity of the transgenic alfalfa lines was significantly increased and their MDA content was notably reduced compared with the control group under the treatment of NaCl. The transgenic lines showed higher capability in scavenging oxygen radicals with lighter NBT staining than the control under salt stress. The transgenic lines showed relative lower water loss rate and electrolyte leakage, but relatively higher Na+ content than the control line under salt stress. The relative expression levels of abiotic-stress-related genes (MsHSP23, MsCOR47, MsATPase, and MsRD2) in three transgenic lines were compared with the control, among them, only the expression of MsCOR47 was up-regulated. Consequently, this study offers a novel perspective for exploring the function of MsNIP2 in improving salt tolerance of alfalfa.

Transcriptome and functional analyses reveal ERF053 from Medicago falcata as key regulator in drought resistances.

Medicago falcata L. is an important legume forage grass with strong drought resistant, which could be utilized as an important gene pool in molecular breed of forage grass. In this study, M. falcata seedlings were treated with 400 mM mannitol to simulate drought stress, and the morphological and physiological changes were investigated, as well as the transcriptome changes of M. falcata seedlings at different treatment time points (0 h, 2 h, 6 h, 12 h, 24 h, 36 h and 48 h). Transcriptome analyses revealed four modules were closely related with drought response in M. falcata by WGCNA analysis, and four ERF transcription factor genes related with drought stress were identified (MfERF053, MfERF9, MfERF034 and MfRAP2.1). Among them, MfERF053 was highly expressed in roots, and MfERF053 protein showed transcriptional activation activity by transient expression in tobacco leaves. Overexpression of MfERF053 in Arabidopsis improved root growth, number of lateral roots and fresh weight under drought, salt stress and exogenous ABA treatments. Transgenic Arabidopsis over-expressing MfERF053 gene grew significantly better than the wild type under both drought stress and salt stress when grown in soil. Taken together, our strategy with transcriptome combined WGCNA analyses identified key transcription factor genes from M. falcata, and the selected MfERF053 gene was verified to be able to enhance drought and salt resistance when over-expressed in Arabidopsis.

CYP3A5 and CYP3A7 genetic polymorphisms affect tacrolimus concentration in pediatric patients with nephrotic range proteinuria.

PURPOSE: The purpose of this study was to investigate the potential impact of CYP3A4, CYP3A5, and CYP3A7 polymorphisms on the concentration and efficacy of tacrolimus in a cohort of pediatric patients with nephrotic range proteinuria. METHODS: Genetic variants including CYP3A5*3 (rs776746), CYP3A4*1G (rs2242480), rs4646437, and CYP3A7 rs2257401 and rs10211 were detected in 70 pediatric patients with nephrotic range proteinuria. The relationships of dose-adjusted trough concentration (C0) of tacrolimus with corresponding genotypes were investigated. RESULTS: The tacrolimus concentration in patients without CYP3A5*3 A allele was 94% higher than those with A allele (90.7 vs 54.2, P = 0.00006). The CYP3A7 rs2257401 was also associated with the concentration of tacrolimus. The C allele carriers had an obviously lower C0 than the non-carriers (62.4 vs 90.7, P = 0.001). In addition, there were significant differences in tacrolimus concentration among CYP3A7 rs10211 G carriers and non-carriers; the latter had an almost twofold C0 of the former (101.8 vs 59.6, P = 0.0004). CONCLUSIONS: Our study demonstrated the associations between CYP3A5*3, CYP3A7 rs2257401 and rs10211, and tacrolimus concentration in pediatric patients with nephrotic range proteinuria. Children with CYP3A5*3 A, CYP3A7 rs2257401 C, and rs10211 G alleles might need a higher dose of tacrolimus.

Monoterpenoid inhibitors of NO production from Paeonia suffruticosa.

Three pairs of monoterpene glycosides (1-6), of which compounds 1, 3, and 4 as new compounds, together with one new monoterpene (7) were obtained from the ethanol extract of Paeonia suffruticosa Andrews. Their structures were determined on the basis of chemical methods and spectral data. On this basis, the spectral characteristics of three pairs of monoterpene glycosides were also discussed. The inhibitory effects of isolated compounds on nitric oxide (NO) production in lipopolysaccharide-activated macrophages were evaluated, and NO production was suppressed significantly by compounds 4-7.

New monoterpene glycosides from Paeonia suffruticosa Andrews and their inhibition on NO production in LPS-induced RAW 264.7 cells.

Nine new monoterpene glycosides (1-9), together with 18 known compounds were obtained from the n-butanol soluble fraction of ethanol extract from Paeonia suffruticosa Andrews. Their structures were determined on the basis of chemical methods and spectroscopic data. The inhibitory effects of these compounds (except compound 6) on nitric oxide (NO) production in lipopolysaccharide-activated macrophages were evaluated. Compounds 1, 21, 23, 25, and 27 showed strong inhibitory activity on NO production with IC(50) values of 31.25, 36.13, 12.41, 6.87, and 41.94 µM, respectively.

Monoterpenes from Paeonia albiflora and their inhibitory activity on nitric oxide production by lipopolysaccharide-activated microglia.

Eleven new monoterpenes, paeonidangenin (1), paeonidanin A (2), paeonidanin B (3), paeonidanin C (4), paeonidanin D (5), paeonidanin E (6), paeoniflorone (7), 4-O-methylbenzoylpaeoniflorin (8), 4-O-methylgalloylpaeoniflorin (9), 4-O-methyldebenzoylpaeoniflorin (10), and 4-O-methylalbiflorin (11), were isolated from the 60% ethanol extract of the roots of Paeonia albiflora. Their structures were determined primarily on the basis of 1D and 2D NMR techniques and MS studies. Paeonidanins D (5) and E (6) are unprecedented examples of "cage-like" monoterpene dimers. The inhibitory effects of the isolated compounds on nitric oxide production by lipopolysaccharide (LPS)-activated N9 microglia were evaluated.

A pair of 3-epimeric physalins from Physalis alkekengi L. var. franchetii.

A pair of new 3-epimeric steroids possessing a rare 13,14-seco-16,24-cycloergostane skeleton, 3alpha-methoxy-2,3-dihydro-4,7-didehydrophysalin B (1), and 3beta-methoxy-2,3-dihydro-4,7-didehydrophysalin B (2) were isolated from the 60% EtOH extract of the stems and leaves of Physalis alkekengi L. var. franchetii. Their structures were characterized on the basis of spectral evidence.

Two new megastigmane glycosides, physanosides A and B, from Physalis alkekengi L. var. franchetii, and their effect on NO release in macrophages.

Two new megastigmane glycosides, physanosides A and B (1 and 2, resp.), were isolated from Physalis alkekengi L. var. franchetii, together with four known compounds (6S,9R)-roseoside (3), (6S,9S)-roseoside (4), (6R,9S)-3-oxo-alpha-ionol beta-D-glucopyranoside (5), and citroside A (6). Their structures were elucidated on the basis of physicochemical evidence, in-depth NMR spectroscopic analysis, high-resolution mass spectrometry, and CD spectroscopy, and their inhibitory effect on NO production was also examined. Compounds 2 and 3 exhibited strong inhibition on lipopolysaccharide-induced NO release by macrophages with IC(50) values of 9.93 and 7.31 microM, respectively.

Steroids and flavonoids from Physalis alkekengi var. franchetii and their inhibitory effects on nitric oxide production.

Four new steroidal compounds possessing a rare 13,14-seco-16,24-cycloergostane skeleton, physalin Y ( 1), physalin Z ( 2), physalin I ( 6), and physalin II ( 7), were isolated from the 80% EtOH extract of calyces from Physalis alkekengi var. franchetii together with 11 known steroids ( 3- 5 and 8- 15) and eight flavonoids ( 16- 23). The structures of the new compounds were elucidated primarily on the basis of 1D and 2D NMR and mass spectroscopic studies. The relative configuration of the 3-OH group of 3, 4, and 5 was also established or re-established. The inhibitory effects of compounds 1- 23 on nitric oxide production in lipopolysaccaride-activated macrophages were evaluated.