Heterologous expression and antimicrobial activity of OsGASR3 from rice (Oryza sativa L.).

According to an in silico analysis, OsGASR3 (LOC_Os03g55290) from rice (Oryza sativa L.) was predicted to be involved in plant defense mechanisms. A semi-quantitative reverse transcription polymerase chain reaction assay revealed that OsGASR3 is highly expressed in the inflorescences of Thai jasmine rice (O. sativa L. subsp. indica 'KDML 105'). To characterize the biological activity of OsGASR3, we produced an OsGASR3-glutathione S-transferase fusion protein in Escherichia coli Rosetta-gami (DE3) cells for a final purified recombinant OsGASR3 yield of 0.65 mg/L. The purified OsGASR3 inhibited the hyphal growth of Fusarium oxysporum f.sp. cubense and Helminthosporium oryzae at a relatively low concentration (7.5 µg/mL). Furthermore, OsGASR3 exhibited in planta inhibitory activity against Xanthomonas campestris, suggesting its involvement in defense mechanisms, in addition to its previously reported functions affecting growth and development. These observations indicate that recombinant OsGASR3 may be useful for protecting agriculturally important crops against pathogenic microbes.

Two novel antimicrobial defensins from rice identified by gene coexpression network analyses.

Defensins form an antimicrobial peptides (AMP) family, and have been widely studied in various plants because of their considerable inhibitory functions. However, their roles in rice (Oryza sativa L.) have not been characterized, even though rice is one of the most important staple crops that is susceptible to damaging infections. Additionally, a previous study identified 598 rice genes encoding cysteine-rich peptides, suggesting there are several uncharacterized AMPs in rice. We performed in silico gene expression and coexpression network analyses of all genes encoding defensin and defensin-like peptides, and determined that OsDEF7 and OsDEF8 are coexpressed with pathogen-responsive genes. Recombinant OsDEF7 and OsDEF8 could form homodimers. They inhibited the growth of the bacteria Xanthomonas oryzae pv. oryzae, X. oryzae pv. oryzicola, and Erwinia carotovora subsp. atroseptica with minimum inhibitory concentration (MIC) ranging from 0.6 to 63µg/mL. However, these OsDEFs are weakly active against the phytopathogenic fungi Helminthosporium oryzae and Fusarium oxysporum f.sp. cubense. This study describes a useful method for identifying potential plant AMPs with biological activities.

Potential of Synechocystis PCC 6803 as a novel cyanobacterial chassis for heterologous expression of enzymes in the trans-resveratrol biosynthetic pathway.

Selected model strains of phototrophic cyanobacteria have been genetically engineered for heterologous expression of numerous enzymes. In the present study, we initially explored the heterologous expression of enzymes involved in trans-resveratrol production, namely, the production of tyrosine ammonia-lyase, coumaroyl CoA-ligase, and stilbene synthase, in the unicellular cyanobacterium Synechocystis PCC 6803. Under the promoters Ptrc1Ocore and Ptrc1O, the respective genes were transcribed and translated into the corresponding soluble proteins at concentrations of 16-34 µg L(-1). The expression levels of these enzymes did not affect the growth rate of the cyanobacterial cells. Interestingly, coumaroyl CoA-ligase expression slightly increased the chlorophyll a content of the cells. Overall, our results suggest that the complete pathway of trans-resveratrol production can be engineered in Synechocystis PCC 6803.