Coronavirus (CoV) envelope (E) protein is a small structural protein critical for virion morphogenesis and release. The recently characterized E protein ion channel activity (EIC) has also been implicated in modulating viral pathogenesis. In this study, we used infectious bronchitis coronavirus (IBV) as a model to study EIC. Two recombinant IBVs (rIBVs) harboring EIC-inactivating mutations -rT16A and rA26F -were serially passaged, and several compensatory mutations were identified in the transmembrane domain (TMD). Two rIBVs harboring these putative EICreverting mutations -rT16A/A26V and rA26F/F14N -were recovered. Compared with the parental rIBV-p65 control, all four EIC mutants exhibited comparable levels of intracellular RNA synthesis, structural protein production, and virion assembly. Our results showed that the IBV EIC contributed to the induction of ER stress response, as up-regulation of ER stress-related genes was markedly reduced in cells infected with the EIC-defective mutants. EIC-defective mutants also formed smaller plaques, released significantly less infectious virions into the culture supernatant, and had lower levels of viral fitness in cell culture. Significantly, all these defective phenotypes were restored in cells infected with the putative EIC revertants. EIC mutations were also implicated in regulating IBV-induced apoptosis, induction of pro-inflammatory cytokines, and viral pathogenicity in vivo. Taken together, this study highlights the importance of CoV EIC in modulating virion release and various aspects of CoV -host interaction.

Pathogens invade intestinal mucosal barrier through phagocytosis of antigen presenting cells (dendritic cell, microfold cells), or through the invasion into the intestinal epithelial directly. Some pathogens could damage the cell junction between epithelial cells and use the paracellular pathway as an entrance to invade. Moreover, some Lactobacillus could inhibit the adhesion of the pathogens and protect the integrity of the cell junction and mucosal barrier. This research focused on the potential therapeutic effect of Lactobacillus fructosus (L. fructosus) C2 to attenuate ETEC K88 or S. typhimurium SL1344 induced changes to mucosal barrier. The results demonstrated that treatment of polarized Caco-2 cells with L. fructosus C2 reduced the permeation of dextran, and expression of IL-8, p-ERK, and p-JNK when cells were infected with pathogenic bacteria. The findings indicated that L. fructosus C2 exerted a protective effect against the damage to the integrity of Caco-2 cells by ETEC or S. typhimurium infection.

Camelina sativa (L.) Crantz is an important Brassicaceae oil crop with a number of excellent agronomic traits including low water and fertilizer input, strong adaptation and resistance. Furthermore, its short life cycle and easy genetic transformation, combined with available data of genome and other "-omics" have enabled camelina as a model oil plant to study lipid metabolism regulation and genetic improvement. Particularly, camelina is capable of rapid metabolic engineering to synthesize and accumulate high levels of unusual fatty acids and modified oils in seeds, which are more stable and environmentally friendly. Such engineered camelina oils have been increasingly used as the super resource for edible oil, health-promoting food and medicine, biofuel oil and high-valued chemical production. In this review, we mainly highlight the latest advance in metabolic engineering towards the predictive manipulation of metabolism for commercial production of desirable bio-based products using camelina as an ideal platform. Moreover, we deeply analysis camelina seed metabolic engineering strategy and its promising achievements by describing the metabolic assembly of biosynthesis pathways for acetyl glycerides, hydroxylated fatty acids, medium-chain fatty acids, w-3 long-chain polyunsaturated fatty acids, palmitoleic acid (w-7) and other high-value oils. Future prospects are discussed, with a focus on the cutting-edge techniques in camelina such as genome editing application, fine directed manipulation of metabolism and future outlook for camelina industry development.

2′-fucosyllactose (2′-FL), one of the simplest but most abundant oligosaccharides in human milk, has been demonstrated to have many positive benefits for the healthy development of newborns. However, the high-cost production and limited availability restrict its widespread use in infant nutrition and further research on its potential functions. In this study, on the basis of previous achievements, we developed a powerful cell factory by using a lacZ-mutant Escherichia coli C41 (DE3)ΔZ to ulteriorly increase 2′-FL production by feeding inexpensive glycerol. Initially, we co-expressed the genes for GDP-L-fucose biosynthesis and heterologous α-1,2-fucosyltransferase in C41(DE3)ΔZ through different plasmid-based expression combinations, functionally constructing a preferred route for 2′-FL biosynthesis. To further boost the carbon flux from GDP-L-fucose toward 2′-FL synthesis, deletion of chromosomal genes (wcaJ, nudD, and nudK) involved in the degradation of the precursors GDP-L-fucose and GDP-mannose were performed. Notably, the co-introduction of two heterologous positive regulators, RcsA and RcsB, was confirmed to be more conducive to GDP-L-fucose formation and thus 2′-FL production. Further a genomic integration of an individual copy of α-1,2-fucosyltransferase gene, as well as the preliminary optimization of fermentation conditions enabled the resulting engineered strain to achieve a high titer and yield. By collectively taking into account the intracellular lactose utilization, GDP-L-fucose availability, and fucosylation activity for 2′-FL production, ultimately a highest titer of 2′-FL in our optimized conditions reached 6.86 g/L with a yield of 0.92 mol/mol from lactose in the batch fermentation. Moreover, the feasibility of mass production was demonstrated in a 50-L fed-batch fermentation system in which a maximum titer of 66.80 g/L 2′-FL was achieved with a yield of 0.89 mol 2′-FL/mol lactose and a productivity of approximately 0.95 g/L/h 2′-FL. As a proof of concept, our preliminary 2′-FL production demonstrated a superior production performance, which will provide a promising candidate process for further industrial production.

As an important oilseed worldwide, Camelina sativa is being increasingly explored for its use in production of food, feed, biofuel and industrial chemicals. However, detailed mechanisms of camelina oil biosynthesis and accumulation, particularly in vegetative tissues, are understood to a very small extent. Here, we present genome-wide identification, cloning and functional analysis of phospholipid diacylglycerol acyltransferase (PDAT) in C. sativa, which catalyses the final acylation step in triacylglycerol (TAG) biosynthesis by transferring a fatty acyl moiety from a phospholipid to diacylglycerol (DAG). We identified five genes (namely CsPDAT1-A, B, and C and CsPDAT2-A and B) encoding PDATs from the camelina genome. CsPDAT1-A is mainly expressed in seeds, whereas CsPDAT1-C preferentially accumulates in flower and leaf tissues. High expression of CsPDAT2-A and CsPDAT2-B was detected in stem and root tissues, respectively. Cold stress induced upregulation of CsPDAT1-A and CsPDAT1-C expression by 3.5- and 2.5-fold, respectively, compared to the control. Salt stress led to an increase in CsPDAT2-B transcripts by 5.1-fold. Drought treatment resulted in an enhancement of CsPDAT2-A mRNAs by twofold and a reduction of CsPDAT2-B expression. Osmotic stress upregulated the expression of CsPDAT1-C by 3.3-fold. Furthermore, the cDNA clones of these CsPDAT genes were isolated for transient expression in tobacco leaves. All five genes showed PDAT enzymatic activity and substantially increased TAG accumulation in the leaves, with CsPDAT1-A showing a higher preference for ɑ-linolenic acid (18:3 ω-3). Overall, this study demonstrated that different members of CsPDAT family contribute to TAG synthesis in different tissues. More importantly, they are involved in different types of stress responses in camelina seedlings, providing new evidence of their roles in oil biosynthesis and regulation in camelina vegetative tissue. The identified CsPDATs may have practical applications in increasing oil accumulation and enhancing stress tolerance in other plants as well.