[Current advance on molecular genetic regulation of rice fertility].

Hybrid rice has contributed greatly to global food security. Cytoplasmic male sterility (CMS) and photo/ thermo sensitive genic male sterility (P/TGMS) are genetic bases for three-line and two-line hybrid rice production, respectively. In contrast, (sub-) specific hybrid sterility (HS) is a major barrier for utilization of hybrid vigor of distant hybrid rice. Therefore, understanding the molecular regulatory mechanism of rice fertility is a key technical issue for hybrid rice industry, and a long-standing basic scientific issue for nuclear-cytoplasmic interaction and reproductive isolation. Chinese geneticists of plant sciences have made tremendous contributions on the molecular genetic basis of rice fertility related to hybrid rice production. Here, we review the development of hybrid rice production systems in China and summarize current advance on genetic basis and molecular mechanism of CMS, P/TGMS, and HS involved in hybrid rice. We also discuss problems of hybrid rice production in China and point out new direction for future utilization of heterosis in rice.

Robust multi-type plasmid modifications based on isothermal in vitro recombination.

A robust strategy for multi-type plasmid modifications is developed based on the isothermal in vitro recombination technology, by which any combination of the sequence modifications can be efficiently achieved in plasmids at any desired position in a seamless manner. As an example, we showed that a plasmid modification with insertion of a GFP gene, deletion of a 623-bp fragment, and substitution of an ampicillin resistance gene by a kanamycin resistance gene was accomplished simultaneously by this method. Therefore, the isothermal in vitro recombination-based multi-type plasmid modification strategy is a useful approach for broad application prospects in molecular biology studies.

The effects of Glycyrrhizae uralenis and its major bioactive components on pharmacokinetics of daphnetin in Cortex daphnes in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Glycyrrhizae uralenis (GU) is often prescribed together with Cortex daphnes (CD) in traditional Chinese medicinal practice to increase the efficacy of CD on the treatment of rheumatoid arthritis (RA), but the reasons were still unknown. In order to clarify the rationality of herbaceous compatibility between CD and GU, the comparative evaluations on pharmacokinetic behaviors of daphnetin (a predominantly active ingredient in CD) after intragastric administration of CD and CD-GU (combination of CD and GU) extract were studied. In addition, the effects of glycyrrhizin and liquiritin, active ingredients of Glycyrrhiza triterpenes and Glycyrrhiza flavones respectively, on the pharmacokinetics of daphnetin were also investigated. MATERIALS AND METHODS: Five groups of rats were orally administered with CD extract, CD-GU extract, pure daphnetin, co-administration of daphnetin and glycyrrhizin as well as co-administration of daphnetin and liquiritin at the same single dose of daphnetin (20 mg/kg). The rat plasma concentrations of daphnetin were determined by our developed UPLC-MS/MS method. The pharmacokinetics of daphnetin in above groups were investigated and compared. RESULTS: Comparing with oral administration of CD extract, AUC and Tmax of daphnetin significantly increased after giving CD-GU (p<0.05). In addition, in comparison to daphnetin alone, co-administration of daphnetin with liquiritin significantly increased the AUC and Cmax of daphnetin for ~1.5-fold, while co-administered with glycyrrhizin showed limited impact on the pharmacokinetics of daphnetin. CONCLUSIONS: In this study, it was found that liquiritin, one of the major components of GU, significantly enhanced the bioavailability of the main component daphnetin in CD. In addition, the bioavailability of daphnetin in the CD-GU prescription was also significantly higher than that in CD alone, which could be due to liquiritin. Such results explained the mechanism of the increased efficacy in treating RA with the combined use of CD and GU.

In vivo antibacterial activity of MRX-I, a new oxazolidinone.

MRX-I is a potent oxazolidinone antibiotic against Gram-positive pathogens, including methicillin-resistant Staphylococcus aureus (MRSA), penicillin-resistant Streptococcus pneumoniae (PRSP), penicillin-intermediate S. pneumoniae (PISP), and vancomycin-resistant enterococci (VRE). In this study, the in vivo efficacy of orally administered MRX-I was evaluated using linezolid as a comparator. MRX-I showed the same or better efficacy than linezolid in both systemic and local infection models against the tested strains.

Characterization of a novel DUF1618 gene family in rice.

Domain of unknown function (DUF) proteins represent a number of gene families that encode functionally uncharacterized proteins in eukaryotes. For example, DUF1618 family members in plants possess a 56-199-amino acid conserved domain and this family has not been described previously. Here, we report the characterization of 121 DUF1618 genes identified in the rice genome. Based on phylogenetic analysis, the rice DUF1618 family was divided into two major groups, each group consisting of two clades. Most DUF1618 genes with close phylogenetic relationships are located in gene clusters on the chromosomes, indicating that gene duplications increased the number of DUF1618 genes. A search for DUF1618 genes in genomic and/or expressed sequence tag databases for 35 other plant species showed that DUF1618 genes are only present in several monocot plants, suggesting that DUF1618 is a new gene family that originated after the dicot-monocot divergence. Based on public microarray databases, most rice DUF1618 genes are expressed at relatively low levels. Further experimental analysis showed that the transcriptional levels of some DUF1618 genes varied in different cultivars, and some responded to stress and hormone conditions, suggesting their important roles for development and fitness in rice (Oryza sativa L.).

The ATP-binding cassette transporter OsABCG15 is required for anther development and pollen fertility in rice.

Plant male reproductive development is a complex biological process, but the underlying mechanism is not well understood. Here, we characterized a rice (Oryza sativa L.) male sterile mutant. Based on map-based cloning and sequence analysis, we identified a 1,459-bp deletion in an adenosine triphosphate (ATP)-binding cassette (ABC) transporter gene, OsABCG15, causing abnormal anthers and male sterility. Therefore, we named this mutant osabcg15. Expression analysis showed that OsABCG15 is expressed specifically in developmental anthers from stage 8 (meiosis II stage) to stage 10 (late microspore stage). Two genes CYP704B2 and WDA1, involved in the biosynthesis of very-long-chain fatty acids for the establishment of the anther cuticle and pollen exine, were downregulated in osabcg15 mutant, suggesting that OsABCG15 may play a key function in the processes related to sporopollenin biosynthesis or sporopollenin transfer from tapetal cells to anther locules. Consistently, histological analysis showed that osabcg15 mutants developed obvious abnormality in postmeiotic tapetum degeneration, leading to rapid degredation of young microspores. The results suggest that OsABCG15 plays a critical role in exine formation and pollen development, similar to the homologous gene of AtABCG26 in Arabidopsis. This work is helpful to understand the regulatory network in rice anther development.

An efficient rice mutagenesis system based on suspension-cultured cells.

Plant mutants are important bio-resources for crop breeding and gene functional studies. Conventional methods for generating mutant libraries by mutagenesis of seeds with physical or chemical agents are of low efficiency. Here, we developed a highly-efficient ethyl methanesulfonate (EMS) mutagenesis system based on suspension-cultured cells, with rice (Oryza sativa L.) as an example. We show that treatment of suspension-cultured tiny cell clusters with 0.4% EMS for 18-22 h followed by differentiation and regeneration produced as high as 29.4% independent mutant lines with visible phenotypic variations, including a number of important agronomic traits such as grain size, panicle size, grain or panicle shape, tiller number and angle, heading date, male sterility, and disease sensitivity. No mosaic mutant was observed in the mutant lines tested. In this mutant library, we obtained a mutant with an abnormally elongated uppermost internode. Sequencing and functional analysis revealed that this is a new allelic mutant of eui (elongated uppermost internode) caused by two point mutations in the first exon of the EUI gene, representing a successful example of this mutagenesis system.

In vitro metabolism in Sprague-Dawley rat liver microsomes of forsythoside A in different compositions of Shuang-Huang-Lian.

Shuang-Huang-Lian (SHL), a traditional Chinese formula containing Lonicerae japonicae flos (LJF), Scutellariae radix (SR) and Forsythiae fructus (FF), is commonly used to treat acute upper respiratory tract infection, acute bronchitis and light pneumonia. Forsythoside A is one of the main active ingredients in Forsythiae fructus, a key herb in SHL. In the present study, effects of different compositions in SHL on the in vitro metabolism in Sprague-Dawley rat liver microsomes of forsythoside A were investigated. The observations from Sprague-Dawley rat liver microsomes in the presence of ß-NADPH or UDPGA that forsythoside A may be the substrates of CYP3A4, CYP2C9, CYP1A2, UGT1A6, UGT1A3, UGT1A1 and UGT1A9; Chlorogenic acid may be the substrates of CYP3A4, CYP2C9, CYP1A2, CYP2C19, UGT1A6, UGT1A3 and UGT1A1; Baicalin may be the substrates of CYP3A4, CYP2C19, CYP1A2, UGT1A9, UGT1A1 and UGT1A3; Baicalein may be the substrates of CYP3A4, CYP2E1 and UGT1A6. It was also found that the residue of forsythoside A in SHL, FF+LJF and FF+SR was greatly increased compared with that in FF in Sprague-Dawley rat liver microsomes in the presence of ß-NADPH or UDPGA, which indicated that the metabolism of forsythoside A in SHL may be influenced by chlorogenic acid in LJF acting on the CYP3A4, CYP2C9, CYP1A2, UGT1A6, UGT1A3 and UGT1A1; baicalin in SR acting on the CYP3A4, CYP1A2, UGT1A9, UGT1A1 and UGT1A3; baicalein acting on the CYP3A4 and UGT1A6 respectively.

Intestinal absorption of forsythoside A in different compositions of Shuang-Huang-Lian.

Shuang-Huang-Lian (SHL), a traditional Chinese formula containing Lonicerae japonicae flos (LJF), Scutellariae radix (SR) and Forsythiae fructus (FF), is commonly used to treat acute upper respiratory tract infection, acute bronchitis and light pneumonia. Forsythoside A is one of the main active ingredients in Forsythiae fructus, a key herb in SHL. In the present study, effects of different compositions in SHL on the intestinal absorption of forsythoside A were investigated. The observations from in situ intestinal circulation model showed that A/%(h(-1)) of forsythoside A in FF+LSF, FF+SR and SHL were all reduced greatly compared with that in FF. However, in pharmacokinetics study, C(max) and AUC(0→1440) of forsythoside A all increased and T(1/2) prolonged in SHL, FF+LJF and FF+SR compared with FF. The results indicated that the different compositions of SHL decreased absorption but increased bioavailability of forsythoside A, which may be related to its metabolism inhibited in intestine or liver.

[Intestinal absorption of forsythoside A by rat circulation in situ].

This study is to investigate the effects of concentration, intestinal section, pH, paracellular route, substrate/inhibitor of enzyme (CYP3A) and proteins (P-gp, MRP2, SGL1) on the absorption of forsythoside A. The absorption of three concentrations (2.6, 5.2, and 10.4 microg x mL(-1)) of forsythoside A in different intestinal segments was studied with phenol red as the marker by rat circulation in situ. The results showed that the residue of forsythoside A with different concentrations had little significant difference from that obtained after perfusing via duodenum, jejunum, ileum and colon, which indicated that the absorption of forsythoside A was passive diffusion and had no difference in different segments of rat intestine. The residue of forsythoside A increased to 466.160 and 463.429 microg respectively when cyclosporine (4 microg x mL(-1)) or midazolam (50 micromol x L(-1)) was added to the circulation fluid, which showed significant difference compared to the control group (P < 0.05). Moreover, the residue of forsythoside A showed a tendency of increase with the increase of cyclosporine or midazolam. When digoxin (50 micromol x L(-1)) or EDTA (10 microg x mL(-1)) was added to the circulation fluid, the residue of forsythoside A decreased to 325.110 and 369.888 microg respectively, which showed significant difference as compared to the control group (P < 0.05). Besides, the residue of forsythoside A showed a tendency of reduction with the increase of digoxin or EDTA. However, there is no significant change in the absorption of forsythoside A when the different concentrations of mannitol were added to the circulation fluid. The results above indicated that the absorption of forsythoside A was mainly passive diffusion and involved paracellular route at the same time. In addition, the substrates of P-gp or CYP3A had dose-dependent effect on the absorption of forsythoside A.