To optimize Agrobacterium-mediated transient transformation assay in mulberry (Morus alba L.), various infiltration methods, Agrobacterium tumefaciens (A. tumefaciens) strains, and bacterial concentrations were tested in mulberry seedlings. Compared with LBA4404, GV3101 harboring pBE2133 plasmids presented stronger GUS signals at 3 days post infiltration using syringe. Recombinant plasmids pBE2133:GFP and pBE2133:GFP:MaFT were successfully constructed. Transient expression of MaFT:GFP protein was found in leaves, petiole (cross section), and shoot apical meristem (SAM) of mulberry according to the GFP signal. Moreover, MaFT:GFP mRNA was also detected in leaves and SAM via RT-PCR and qRT-PCR. An efficient transient transformation system could be achieved in mulberry seedlings by syringe using A. tumefaciens GV3101 at the OD₆₀₀ of 0.5. The movement of MaFT expression from leaves to SAM might trigger the precocious flowering of mulberry. Graphical abstract Transient MaFT expression could be achieved in mulberry by syringe using GV3101 (OD₆₀₀ = 0.5). MaFT might trigger the precocious flowering of mulberry.

Mulberry (Morus alba L.) has been an economically important food crop for the domesticated silkworm, Bombyx mori, in China for more than 5000 years. However, little is known about the mechanism underlying mulberry response to environmental stress. In this study, quantitative proteomics was applied to elucidate the molecular mechanism of drought response in mulberry. A total of 604 differentially expressed proteins (DEPs) were identified via LC-MS/MS. The proteomic profiles associated with antioxidant enzymes, especially five glutathione peroxidase (GPX) isoforms, as a scavenger of reactive oxygen species (ROS), were systematically increased in the drought-stressed mulberry. This was further confirmed by gene expression and enzymatic activity. Furthermore, overexpression of the GPX isoforms led to enhancements in both antioxidant system and ROS-scavenging capacity, and greater tolerance to drought stress in transgenic plants. Taken together, these results indicated that GPX-based antioxidant enzymes play an important role in modulating mulberry response to drought stress, and higher levels of GPX can improve drought tolerance through enhancing the capacity of the antioxidant system for ROS scavenging.

The oligosaccharide 3-deoxy-d-manno-oct-2-ulosonic acid (Kdo) is a key component of lipopolysaccharide in Gram-negative bacteria, and is also part of the pectic polysaccharide rhamnogalacturonan (RG-II) of the plant cell wall. The enzyme KDO-8-phosphate synthase (KDO8Ps), encoded by the 2-dehydro-3-deoxyphosphooctonate aldolase (KdsA) gene, catalyzes the first step in the synthesis of Kdo. In this study, the complete coding sequence of the KdsA gene from mulberry leaves was cloned and the primary structure of KDO8Ps was deduced. Alignment of the amino acid sequence of KDO8Ps from mulberry with those of five other plant species revealed a high level of evolutionary conservation. A phylogenetic tree analysis demonstrated a short genetic distance among KDO8Ps proteins of different species. Expression of the KdsA gene was higher in the second leaves than in the eighth leaves of mulberry, and was down-regulated under conditions of high salt or drought stress. Our results suggest that KdsA expression is important for the growth of new plant tissues, and is sensitive to harsh environments.

Autophagy is associated with multiple biological processes and has protective and defensive functions with respect to immunity, inflammation, and resistance to microbial infection. In this experiment, we wished to investigate whether autophagy is a factor in the midgut cell response of Bombyx mori to infection by the B. mori cytoplasmic polyhedrosis virus (BmCPV). Our results indicated that the expression of three autophagy-related genes (BmAtg8, BmAtg5, and BmAtg7) in the midgut did not change greatly after BmCPV infection in B. mori. Basal ATG8/ATG8PE protein expression was detected in different B. mori tissues by using western blot analysis. Immunohistochemistry showed that the ATG8/ATG8PE proteins were located mainly in the cytoplasm. ATG8/ATG8PE protein levels decreased at 12 and 16 h after BmCPV infection. Our results indicate that autophagy responded slightly to BmCPV infection, but could not prevent the invasion and replication of the virus.