Abstract
It is well known that exogenous trehalose can improve resistances of plants to some abiotic and biotic stresses. Nonetheless, information respecting the molecular responses of tobacco leaves to Tre treatment is limited. Here we show that exogenous Tre can rapidly reduce stomatal aperture, up-regulate NADPH oxidase genes and increase O2•-andH2O2 on tobacco leaves at 2 h after treatment. We further demonstrated that imidazole and DPI, inhibitors of NADPH oxidase, can promote recovery of stomatal aperture of tobacco leaves upon trehalose treatment. Exogenous trehalose increased tobacco leaf resistance to tobacco mosaic disease significantly in a concentration-dependent way. To elucidate the molecular mechanisms in response to exogenous trehalose, the transcriptomic responses of tobacco leaves with 10 (low concentration) or 50 (high concentration) mM of trehalose treatment at 2 or 24h were investigated through RNA-seq approach. In total, 1288 differentially expressed genes (DEGs) were found with different conditions of trehalose treatments relative to control. Among them, 1075 (83.5%) were triggered by low concentration of trehalose (10mM), indicating that low concentration of Tre is a better elicitor. Functional annotations with KEGG pathway analysis revealed that the DEGs are involved in metabolic pathway, biosynthesis of secondary metabolites, plant hormone signal transduction, plant-pathogen interaction, protein processing in ER, flavonoid synthesis and circadian rhythm and so on. The protein-protein interaction networks generated from the core DEGs regulated by all conditions strikingly revealed that eight proteins, including ClpB1, HSP70, DnaJB1-like protein, universal stress protein (USP) A-like protein, two FTSH6 proteins, GolS1-like protein and chloroplastics HSP, play a core role in responses to exogenous trehalose in tobacco leaves. Our data suggest that trehalose triggers a signal transduction pathway which involves calcium and ROS-mediated signalings. These core components could lead to partial resistance or tolerance to abiotic and biotic stresses. Moreover, 19 DEGs were chosen for analysis of quantitative real-time polymerase chain reaction (qRT-PCR). The qRT-PCR for the 19 candidate genes coincided with the DEGs identified via the RNA-seq analysis, sustaining the reliability of our RNA-seq data.
Highlights
Trehalose (Tre), as a non-reducing disaccharide, is formed by two α-glucose units linked through α, α-1,1-glucosidic bond (α-D-glucopyranosyl-[1,1]-α-D-glucopyranoside)
Combined with the effect of Tre concentration on inhibition rate of TMV infection, a series of physiological phenotypes of tobacco leaves in response to exogenous Tre were characterized to identify the optimal time-points of Tre treatment for RNA-seq library construction
The sixth and seventh fully expanded true leaves of N. tabaccum at the 8 leaves stage were smeared with 2 ml of 0, 10 and 50 mM of Tre on both sides, respectively, and the ROS species, O2- and H2O2, were monitored at 2h and 24 h after pretreatment separately as described in methods
Summary
Trehalose (Tre), as a non-reducing disaccharide, is formed by two α-glucose units linked through α, α-1,1-glucosidic bond (α-D-glucopyranosyl-[1,1]-α-D-glucopyranoside). Tre biosynthesis and signaling in vivo have been investigated extensively in many different organisms, including bacteria, yeast, fungi, insects, plants and animals[1, 2]. Even though alternative pathways exist in different organisms, biosynthesis of Tre typically includes two steps. T6P, as an intermediate metabolite of Tre biosynthesis, has been proved to function as a sensor for in vivo available sucrose, by this means regulating the responses of organism to the diverse environmental changes directly, which is reasonable as the components of Tre biosynthesis pathway, such as T6P, trehalose and their biosynthetic enzymes are part of an interactive correlation network including sugar and hormone signaling pathways, etc[7]. The components of Tre biosynthesis pathway influence growth and development, and get involved in responses of both abiotic and biotic stresses[7,8,9]. Tre accumulated in Tripogonloliiformis can regulate autophagy that might further confer the plant desiccation tolerance[14]
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