A rice seed-specific glycine-rich protein OsDOR1 interacts with GID1 to repress GA signaling and regulates seed dormancy.

Seed dormancy is an important agronomic trait under the control of complex genetic and environmental interactions, which have not been yet comprehensively understood. From the field screening of rice mutant library generated by a Ds transposable element, we identified a pre-harvest sprouting (PHS) mutant dor1. This mutant has a single insertion of Ds element at the second exon of OsDOR1 (LOC_Os03g20770), which encodes a novel seed-specific glycine-rich protein. This gene successfully complemented the PHS phenotype of dor1 mutant and its ectopic expression enhanced seed dormancy. Here, we demonstrated that OsDOR1 protein binds to the GA receptor protein, OsGID1 in rice protoplasts, and interrupts with the formation OsGID1-OsSLR1 complex in yeast cells. Co-expression of OsDOR1 with OsGID1 in rice protoplasts attenuated the GA-dependent degradation of OsSLR1, the key repressor of GA signaling. We showed the endogenous OsSLR1 protein level in the dor1 mutant seeds is significantly lower than that of wild type. The dor1 mutant featured a hypersensitive GA-response of α-amylase gene expression during seed germination. Based on these findings, we suggest that OsDOR1 is a novel negative player of GA signaling operated in the maintenance of seed dormancy. Our findings provide a novel source of PHS resistance.

Ruminococcus gnavus ameliorates atopic dermatitis by enhancing Treg cell and metabolites in BALB/c mice.

BACKGROUND: Ruminococcus gnavus (R. gnavus) are mucin-degrading gut bacteria that play a key role in the early colonization of the gut by serving as endogenous sources of nutrients. They can also influence immune development. We had previously reported a lower abundance of R. gnavus in infants with atopic dermatitis (AD) compared with that in healthy subjects. However, the underlying mechanisms remain unclear. In this study, we investigated the effect of orally administered R. gnavus on antibiotic treatment-induced gut dysbiosis (and the underlying mechanism) in a mouse model of AD. METHODS: Four-week-old female BALB/C mice were administered antibiotic cocktails for 2 weeks. R. gnavus was orally administered throughout the study duration. At 6 weeks of age, AD was induced by epidermal sensitization with ovalbumin. AD phenotypes and systemic and gut immune responses were investigated. RESULTS: Orally administered R. gnavus significantly reduced AD-associated parameters (i.e., transepidermal water loss, clinical score, total serum immunoglobulin (Ig) E level, OVA-specific IgE level, and skin inflammation). R. gnavus treatment also resulted in significant downregulation of T helper 2-related cytokine mRNA and upregulation of interleukin (IL)-10 and Foxp3 in the skin. The population of CD4+ FOXP3+ T cells in mesenteric- and skin-draining lymph nodes and butyrate levels in the cecum increased in R. gnavus-administered AD mice. CONCLUSIONS: Immune modulation by orally administered R. gnavus may alleviate AD symptoms through the enhancement of regulatory T-cell counts and short-chain fatty acids production in AD mice.

CCOAOMT1, a candidate cargo secreted via VAMP721/722 secretory vesicles in Arabidopsis.

We previously found that VAMP721/722 SNARE proteins guide secretory vesicles to pathogen-attacking sites during immune responses in Arabidopsis, which suggests that these vesicles should deliver immune molecules. However, the lethality of vamp721 vamp722 double null mutant makes it difficult to understand the nature of cargo transported via VAMP721/722 vesicles. Since VAMP721/722-depleted (VAMP721+/-VAMP722-/- and VAMP721-/-VAMP722+/-) plants show compromised resistance to extracellular pathogens, we assume that an immune protein secreted through the VAMP721/722-engaged exocytosis would be remained more in VAMP721/722-depleted plants than WT. By comparing intracellular proteins between WT and VAMP721/722-depleted plants, we found caffeoyl-CoA O-methyltransferase 1 (CCOAOMT1) involved in the lignin biosynthesis was more abundantly detected in both VAMP721/722-depleted lines than WT. Plants are well-known to deposit secondary cell walls as physical barriers at pathogen-attempting sites. Therefore, extracellular detection of CCOAOMT1 and impaired resistance to Pseudomonas syringae DC3000 in ccoaomt1 plants suggest that plants secrete cell wall-modifying enzymes at least including CCOAOMT1 to reinforce the secondary cell walls for immunity.

Elevated aldolase 1A, retrogene 1 expression induces cardiac apoptosis in rat experimental autoimmune myocarditis model.

Acute myocarditis is an unpredictable heart disease that is caused by inflammation-associated cell death. Although viral infection and drug exposure are known to induce acute myocarditis, the molecular basis for its development remains undefined. Using proteomics and molecular analyses in myosin-induced rat experimental autoimmune myocarditis (EAM), we identified that elevated expression of aldolase 1A, retrogene 1 (Aldoart1) is critical to induce mitochondrial dysfunction and acute myocarditis development. Here, we demonstrate that cardiac cell death is associated with increased expressions of proapoptotic genes in addition to high levels of glucose, lactate, and triglyceride in metabolite profiling. The functional protein association network analysis also suggests that Aldoart1 upregulation correlates with high levels of dihydroxyacetone kinase and triglyceride. In H9c2 cardiac cells, lipopolysaccharides (LPS) or high glucose exposure significantly increases the cytochrome c release and the conversion of pro-caspase 3 into the cleaved form of caspase 3. We also found that LPS- or glucose-induced toxicities are almost completely reversed by siRNA-mediated knockdown of Aldoartl, which consequently increases cell viability. Together, our study strongly suggests that Aldoart1 may be involved in inducing mitochondrial apoptotic processes and can be a novel therapeutic target to prevent the onset of acute myocarditis or cardiac apoptosis.

Clinical and biochemical relevance of monounsaturated fatty acid metabolism targeting strategy for cancer stem cell elimination in colon cancer.

Lipid metabolism is associated with colon cancer prognosis and incidence. Stearoyl-CoA desaturase 1 (SCD1), which converts fully saturated fatty acids (SFAs) to monounsaturated fatty acids (MUFAs), has been suggested as a vulnerable target for selective elimination of cancer stem cells (CSCs). However, the clinical significance and physiological role of SCD1 in CSCs has not been well demonstrated. Here, we showed the clinical and biochemical relevance of blocking SCD1 to target CSCs by analyzing human colon cancer data from TCGA and through lipidomic profiling of CSCs with or without SCD1 inhibition using mass spectrometry. Positive associations between SCD1 expression and colorectal cancer patient clinical status and the expression of CSC-related genes (WNT and NOTCH signaling) were found based on TCGA data analysis. Lipidomic profiling of CSCs and bulk cancer cells (BCCs) using mass spectrometry revealed that colon CSCs contained a distinctive lipid profile, with higher free MUFA and lower free SFA levels than in BCCs, suggesting that enhanced SCD1 activity generates MUFAs that may support WNT signaling in CSCs. In addition, all identified phosphatidyl-ethanolamine-containing MUFAs were found at higher levels in CSCs. Interestingly, we observed lower phosphatidyl-serine (18:1/18:0), phosphatidyl-choline (PC; p-18:0/18:1)), and sphingomyelin (SM; d18:1/20:0 or d16:1/22:0) levels in CSCs than in BCCs. Of those, SCD1 inhibition, which efficiently diminished free MUFA levels, increased those specific PC and SM and MUFAs in CSCs promptly. These results suggest that these specific lipid composition is critical for CSC stem cell maintenance. In addition, not only free MUFAs, which are known to be required for WNT signaling, but also other phospholipids, such as SM, which are important for lipid raft formation, may mediate other cell signaling pathways that support CSC maintenance. Comparison of the lipidomic profiles of colon cancer cells with those of previously reported for glioma cells further demonstrated the tissue specific characteristics of lipid metabolism in CSCs.

Loss-of-function screens of druggable targetome against cancer stem-like cells.

Cancer stem-like cells (CSLCs) contribute to the initiation and recurrence of tumors and to their resistance to conventional therapies. In this study, small interfering RNA (siRNA)-based screening of ∼4800 druggable genes in 3-dimensional CSLC cultures in comparison to 2-dimensional bulk cultures of U87 glioma cells revealed 3 groups of genes essential for the following: survival of the CSLC population only, bulk-cultured population only, or both populations. While diverse biologic processes were associated with siRNAs reducing the bulk-cultured population, CSLC-eliminating siRNAs were enriched in a few functional categories, such as lipid metabolism, protein metabolism, and gene expression. Interestingly, siRNAs that selectively reduced CSLC only were found to target genes for cholesterol and unsaturated fatty acid synthesis. The lipidomic profile of CSLCs revealed increased levels of monounsaturated lipids. Pharmacologic blockage of these target pathways reduced CSLCs, and this effect was eliminated by addition of downstream metabolite products. The present CSLC-sensitive target categories provide a useful resource that can be exploited for the selective elimination of CSLCs.-Song, M., Lee, H., Nam, M.-H., Jeong, E., Kim, S., Hong, Y., Kim, N., Yim, H. Y., Yoo, Y.-J., Kim, J. S., Kim, J.-S., Cho, Y.-Y., Mills, G. B., Kim, W.-Y., Yoon, S. Loss-of-function screens of druggable targetome against cancer stem-like cells.

Amelioration of obesity-related characteristics by a probiotic formulation in a high-fat diet-induced obese rat model.

PURPOSE: Obesity is a major public health concern. Despite its multi-factorial etiology, alterations in intestinal microbiota and the immune system are frequently observed. We investigated the effect of Duolac Gold (DG), a probiotic formulation containing 2 Lactobacillus strains (L. acidophilus LA1 and L. rharmnosus LR5), 3 Bifidobacterium (B. bifidum BF3, B. lactis BL3, and B. longum BG7), and Streptococcus thermophilus ST3, on morphometric and metabolic parameters, intestinal microbiota, and intestinal immune responses in a high-fat diet (HFD)-induced obese rat model. METHODS: Rats received either a conventional balanced diet or HFD with or without water containing DG for 8 weeks. HFD-induced adiposity, intestinal microbiota, and changes in inflammatory cytokine, chemokine, and metabolite levels in serum were evaluated. RESULTS: DG administration effectively decreased HFD-induced body weight and modulated morphometric and metabolic parameters. Quantitative analysis of fecal microbiota showed that obese rats given DG exhibited significantly increased levels of Bacteroidetes, Lactobacillus, and Bifidobacterium, with significant decreases in the level of Firmicutes. Serum levels of the inflammatory cytokines and the chemokine were also altered. Serum metabolite analysis revealed that DG administration modulated HFD-induced changes in serum metabolites, including fatty acids (FA), lysophosphatidylcholine, lysophosphatidylethanolamine, phosphatidylcholine (PC), and triacylglycerol (TAG). CONCLUSIONS: DG administration appears to have the potential to alleviate HDF-induced obesity through the modulation of intestinal microbiota, immune responses, and host metabolism, which supports the use of probiotics to treat obesity.

Network analysis of the metabolome and transcriptome reveals novel regulation of potato pigmentation.

To gain insights into the regulatory networks related to anthocyanin biosynthesis and identify key regulatory genes, we performed an integrated analysis of the transcriptome and metabolome in sprouts germinated from three colored potato cultivars: light-red Hongyoung, dark-purple Jayoung, and white Atlantic. We investigated transcriptional and metabolic changes using statistical analyses and gene-metabolite correlation networks. Transcript and metabolite profiles were generated through high-throughput RNA-sequencing data analysis and ultraperformance liquid chromatography quadrupole time-of-flight tandem mass spectrometry, respectively. The identification and quantification of changes in anthocyanin were performed using molecular formula-based mass accuracy and specific features of their MS(2) spectra. Correlation tests of anthocyanin contents and transcriptional changes showed 823 strong correlations (correlation coefficient, R (2)>0.9) between 22 compounds and 119 transcripts categorized into flavonoid metabolism, hormones, transcriptional regulation, and signaling. The connection network of anthocyanins and genes showed a regulatory system involved in the pigmentation of light-red Hongyoung and dark-purple Jayoung potatoes, suggesting that this systemic approach is powerful for investigations into novel genes that are potential targets for the breeding of new valuable potato cultivars.