An S-domain receptor-like kinase, OsESG1, regulates early crown root development and drought resistance in rice.

Plant receptor-like kinase (RLKs) are serine/threonine protein kinases that play fundamental roles in development, innate immunity, and abiotic stress response. Here, we identified an S-domain receptor-like kinase OsESG1 from rice (Oryza sativa), and identified its involvement in early crown root (CR) development and drought response. The OsESG1 kinase domain possessed auto-phosphorylation activity and was able to phosphorylate MBP and His proteins. OsESG1 was expressed ubiquitously in all tissues that were examined, with relatively higher expression in the embryo. And it could be induced to express by treating with PEG, NaCl and ABA. Transgenic plants carrying anti-sense (AS) OsESG1 were generated by knockdown of OsESG1 expression. At the early seedling stage, AS lines had fewer CRs and shorter shoot compared with wild type (WT) plants. IAA flux and the genes' expressions of the auxin responsive and efflux carrier were infected in the AS lines. These results indicated that auxin signaling and polar auxin transport (PAT) were disrupted. The AS lines were more sensitive to osmotic stress compared to WT, and showed excessive accumulation of reactive oxygen species (ROS) and malondialdehyde (MDA), lower activities of antioxidant enzymes, and impaired expressions of stress-related genes under PEG treatment. Results above suggested that OsESG1 may regulate CR initiation and development by controlling auxin response and distribution, and participate in stress response by regulating the activities of antioxidants and expressions of stress-regulated genes.

Evaluating the Effect of Expressing a Peanut Resveratrol Synthase Gene in Rice.

Resveratrol (Res) is a type of natural plant stilbenes and phytoalexins that only exists in a few plant species. Studies have shown that the Res could be biosynthesized and accumulated within plants, once the complete metabolic pathway and related enzymes, such as the key enzyme resveratrol synthase (RS), existed. In this study, a RS gene named PNRS1 was cloned from the peanut, and the activity was confirmed in E. coli. Using transgenic approach, the PNRS1 transgenic rice was obtained. In T3 generation, the Res production and accumulation were further detected by HPLC. Our data revealed that compared to the wild type rice which trans-resveratrol was undetectable, in transgenic rice, the trans-resveratrol could be synthesized and achieved up to 0.697 µg/g FW in seedlings and 3.053 µg/g DW in seeds. Furthermore, the concentration of trans-resveratrol in transgenic rice seedlings could be induced up to eight or four-fold higher by ultraviolet (UV-C) or dark, respectively. Simultaneously, the endogenous increased of Res also showed the advantages in protecting the host plant from UV-C caused damage or dark-induced senescence. Our data indicated that Res was involved in host-defense responses against environmental stresses in transgenic rice. Here the results describes the processes of a peanut resveratrol synthase gene transformed into rice, and the detection of trans-resveratrol in transgenic rice, and the role of trans-resveratrol as a phytoalexin in transgenic rice when treated by UV-C and dark. These findings present new outcomes of transgenic approaches for functional genes and their corresponding physiological functions, and shed some light on broadening available resources of Res, nutritional improvement of crops, and new variety cultivation by genetic engineering.

Overexpression of peanut diacylglycerol acyltransferase 2 in Escherichia coli.

Diacylglycerol acyltransferase (DGAT) is the rate-limiting enzyme in triacylglycerol biosynthesis in eukaryotic organisms. Triacylglycerols are important energy-storage oils in plants such as peanuts, soybeans and rape. In this study, Arachis hypogaea type 2 DGAT (AhDGAT2) genes were cloned from the peanut cultivar 'Luhua 14' using a homologous gene sequence method and rapid amplification of cDNA ends. To understand the role of AhDGAT2 in triacylglycerol biosynthesis, two AhDGAT2 nucleotide sequences that differed by three amino acids were expressed as glutathione S-transferase (GST) fusion proteins in Escherichia coli Rosetta (DE3). Following IPTG induction, the isozymes (AhDGAT2a and AhDGAT2b) were expressed as 64.5 kDa GST fusion proteins. Both AhDGAT2a and AhDGAT2b occurred in the host cell cytoplasm and inclusion bodies, with larger amounts in the inclusion bodies. Overexpression of AhDGATs depressed the host cell growth rates relative to non-transformed cells, but cells harboring empty-vector, AhDGAT2a-GST, or AhDGAT2b-GST exhibited no obvious growth rate differences. Interestingly, induction of AhDGAT2a-GST and AhDGAT2b-GST proteins increased the sizes of the host cells by 2.4-2.5 times that of the controls (post-IPTG induction). The total fatty acid (FA) levels of the AhDGAT2a-GST and AhDGAT2a-GST transformants, as well as levels of C12:0, C14:0, C16:0, C16:1, C18:1n9c and C18:3n3 FAs, increased markedly, whereas C15:0 and C21:0 levels were lower than in non-transformed cells or those containing empty-vectors. In addition, the levels of some FAs differed between the two transformant strains, indicating that the two isozymes might have different functions in peanuts. This is the first time that a full-length recombinant peanut DGAT2 has been produced in a bacterial expression system and the first analysis of its effects on the content and composition of fatty acids in E. coli. Our results indicate that AhDGAT2 is a strong candidate gene for efficient FA production in E. coli.

[Effects of early growth stage shading on rice flag leaf physiological characters and grain growth at grain-filling stage].

In a pot experiment, rice plants were shaded during the period from transplanting to booting, aimed to study the effects of early growth stage shading on the rice growth at grain-filling stage. Comparing with the control, early growth stage shading decreased the tiller number by 26.72%, but increased the flag leaf area and soluble sugar content by 33.86% and 30.23%, respectively. The filled-grain number per panicle, 1000-grain mass, ultimate brown rice mass, and maximum and average grain-filling rates decreased by 8.65%, 4.81%, 9.74%, 20.22%, and 19.13%, and the effective panicle number and grain yield declined by 25.26% and 39.56%, respectively. The peak time of grain-filling rate (Tm) advanced 1.66 days, while the grain-filling time (T99) prolonged 6.80 days. For shading-tolerance variety, its flag leaf Chl a, Chl b, and Chl (a + b) contents at early and mid grain-filling stages, and the protein N and soluble sugar contents and Chl a/b in its flag leaves at grain-filling stage all increased under early growth stage shading, and the ultimate brown rice mass and 1000-grain mass maintained at the similar levels as the control. Consequently, its grain yield reduction rate was lower than that of shading-sensitive variety.