Tandem mass tag-based proteomic analysis of endoplasmic reticulum proteins in mulberry leaves under ultraviolet-B and dark stress.

Mulberry leaves have been used in traditional Chinese medicine due to their antioxidant, antidiabetic, and antihyperlipidemic properties. A previous study showed that ultraviolet-B radiation followed by dark incubation could improve the contents of active ingredients in mulberry leaves, such as moracin N and chalcomoracin. The endoplasmic reticulum (ER) serves as a protein quality control center and the location for protein synthesis, which is involved in the response to the environmental stress in plants. To investigate the mechanisms in response to ultraviolet-B radiation followed by dark incubation (UV + D), ER proteomics was performed on mulberry leaves. The ER protein markers, glucose-regulated protein (GRP78), and calnexin (CNX), were significantly higher in the ER fraction than in the total protein fraction, indicating that the ER was purified. Compared to the control, the abundance of protein disulfide isomerase, UDP-glucose glycoprotein glucosyltransferase, CNX, and calreticulin proteins decreased, while of the abundance of heat shock-related proteins increased under stress. P450 enzyme system-related proteins and ribosomal proteins showed significant increases. These results suggest that under UV + D stress, mulberry leaves activated the cell redox and ER quality control systems, enhancing protein synthesis and weakening N-glycan biosynthesis in the ER to resist the damage.

Integrative omics of Lonicera japonica Thunb. Flower development unravels molecular changes regulating secondary metabolites.

Lonicera japonica Thunb. is an important medicinal plant. The secondary metabolites in L. japonica are diverse and vary in levels during development, leading to the ambiguous evaluation for its medical value. In order to reveal the regulatory mechanism of secondary metabolites during the flowering stages, transcriptomic, proteomic, and metabolomic analyses were performed. The integration analysis of omic-data illustrated that the metabolic changes over the flower developmental stages were mainly involved in sugar metabolism, lipopolysaccharide biosynthesis, carbon conversion, and secondary metabolism. Further proteomic analysis revealed that uniquely identified proteins were mainly involved in glycolysis/phenylpropanoids and tricarboxylic acid cycle/terpenoid backbone pathways in early and late stages, respectively. Transketolase was commonly identified in the 5 developmental stages and 2-fold increase in gold flowering stage compared with juvenile bud stage. Simple phenylpropanoids/flavonoids and 1-deoxy-D-xylulose-5-phosphate were accumulated in early stages and upregulated in late stages, respectively. These results indicate that phenylpropanoids were accumulated attributing to the activated glycolysis process in the early stages, while the terpenoids biosynthetic pathways might be promoted by the transketolase-contained regulatory circuit in the late stages of L. japonica flower development. BIOLOGICAL SIGNIFICANCE: Lonicera japonica Thunb. is a native species in the East Asian and used in traditional Chinese medicine. In order to reveal the regulatory mechanism of secondary metabolites during the flowering stages, transcriptomic, proteomic, and metabolomic analyses were performed. The integration analysis of omic-data illustrated that the metabolic changes over the flower developmental stages were mainly involved in sugar metabolism, lipopolysaccharide biosynthesis, carbon conversion, and secondary metabolism. Our results indicate that phenylpropanoids were accumulated attributing to the activated glycolysis process in the early stages, while the terpenoids biosynthetic pathways might be promoted by the transketolase-contained regulatory circuit in the late stages of L. japonica flower development.

Phosphoproteomics Reveals the Biosynthesis of Secondary Metabolites in Catharanthus roseus under Ultraviolet-B Radiation.

Ultraviolet (UV)-B radiation acts as an elicitor to enhance the production of secondary metabolites in medicinal plants. To investigate the mechanisms, which lead to secondary metabolites in Catharanthus roseus under UVB radiation, a phosphoproteomic technique was used. ATP content increased in the leaves of C. roseus under UVB radiation. Phosphoproteins related to calcium such as calmodulin, calcium-dependent kinase, and heat shock proteins increased. Phosphoproteins related to protein synthesis/modification/degradation and signaling intensively changed. Metabolomic analysis indicated that the metabolites classified with pentoses, aromatic amino acids, and phenylpropanoids accumulated under UVB radiation. Phosphoproteomic and immunoblot analyses indicated that proteins related to glycolysis and the reactive-oxygen species scavenging system were changed under UVB radiation. These results suggest that UVB radiation activates the calcium-related pathway and reactive-oxygen species scavenging system in C. roseus. These changes lead to the upregulation of proteins, which are responsible for the redox reactions in secondary metabolism and are important for the accumulation of secondary metabolites in C. roseus under UVB radiation.