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Plant root border cells (RBCs) prevent the colonization of plant growth‐promoting rhizobacteria (PGPR) at the root tip, rendering the PGPR unable to effectively control pathogens infecting the root tip. In this study, we engineered four strains of Pseudomonas sp. UW4, a typical PGPR strain, each carrying an enhanced green fluorescent protein (EGFP)‐expressing plasmid. The UW4E strain harboured only the plasmid, whereas the UW4E‐flg22 strain expressed a secreted EGFP‐Flg22 fusion protein, the UW4E‐Flg(flg22) strain expressed a non‐secreted Flg22, and the UW4E‐flg22‐D strain expressed a secreted Flg22‐DNase fusion protein. UW4E‐flg22 and UW4E‐flg22‐D, which secreted Flg22, induced an immune response in wheat RBCs and colonized wheat root tips, whereas the other strains, which did not secrete Flg22, failed to elicit this response and did not colonize wheat root tips. The immune response revealed that wheat RBCs synthesized mucilage, extracellular DNA, and reactive oxygen species. Furthermore, the Flg22‐secreting strains showed a 33.8%–93.8% higher colonization of wheat root tips and reduced the root rot incidence caused by Rhizoctonia solani and Fusarium pseudograminearum by 24.6%–35.7% compared to the non‐Flg22‐secreting strains in pot trials. There was a negative correlation between the incidence of wheat root rot and colonization of wheat root tips by these strains. In contrast, wheat root length and dry weight were positively correlated with the colonization of wheat root tips by these strains. These results demonstrate that engineered secretion of Flg22 by PGPR is an effective strategy for controlling root rot and improving plant growth. [ABSTRACT FROM AUTHOR]

The 1-aminocyclopropane-1-carboxylic acid (ACC) pathway that synthesizes ethylene is shared in seed plants, fungi and probably other organisms. However, the evolutionary relationship of the key enzyme ACC oxidase (ACO) in the pathway among organisms remains unknown. Herein, we cloned, expressed and characterized five ACOs from the straw mushroom (Volvariella volvacea) and the oyster mushroom (Pleurotus ostreatus): VvACO1-4 and PoACO. The five mushroom ACOs and the previously identified AbACO of the button mushroom contained all three conserved residues that bound to Fe(II) in plant ACOs. They also had variable residues that were conserved and bound to ascorbate and bicarbonate in plant ACOs and harbored only 1–2 of the five conserved ACO motifs in plant ACOs. Particularly, VvACO2 and AbACO had only one ACO motif 2. Additionally, VvACO4 shared 44.23% sequence identity with the cyanobacterium Hapalosiphon putative functional ACO. Phylogenetic analysis showed that the functional ACOs of monocotyledonous and dicotyledonous plants co-occurred in Type I, Type II and Type III, while putative functional gymnosperm ACOs also appeared in Type III. The putative functional bacterial ACO, functional fungi and slime mold ACOs were clustered in ancestral Type IV. These results indicate that ACO motif 2, ACC and Fe(II) are essential for ACO activity. The ACOs of the other organisms may come from the horizontal transfer of fungal ACOs, which were found ordinarily in basidiomycetes. It is mostly the first case for the horizontal gene transfers from fungi to seed plants. The horizontal transfer of ACOs from fungi to plants probably facilitates the fungal-plant symbioses, plant–land colonization and further evolution to form seeds. [ABSTRACT FROM AUTHOR]

Mushroom β-glucans are potent immunological stimulators in medicine, but their productivities are very low. In this study, we successfully improved its production by promoter engineering in Pleurotus ostreatus. The promoter for β-1,3-glucan synthase gene (GLS) was replaced by the promoter of glyceraldehyde-3-phosphate dehydrogenase gene of Aspergillus nidulans. The homologous recombination fragment for swapping GLS promoter comprised five segments, which were fused by two rounds of combined touchdown PCR and overlap extension PCR (TD-OE PCR), and was introduced into P. ostreatus through PEG/CaCl2-mediated protoplast transformation. The transformants exhibited one to three fold higher transcription of GLS gene and produced 32% to 131% higher yield of β-glucans than the wild type. The polysaccharide yields had a significant positive correlation to the GLS gene expression. The infrared spectra of the polysaccharides all displayed the typical absorption peaks of β-glucans. This is the first report of successful swapping of promoters in filamentous fungi. [ABSTRACT FROM AUTHOR]