Tobacco salicylic acid glucosyltransferase is active toward tuberonic acid (12-hydroxyjasmonic acid) and is induced by mechanical wounding stress

Recently we reported that rice salicylic acid (SA) glucosyltransferase (OsSGT) is active toward 12-hydroxyjasmonic acid (tuberonic acid, TA) and that OsSGT gene expression is induced by wounding stress. Here we report that tobacco SA glucosyltransferase (NtSGT), which is thought to be an ortholog of OsSGT, is also active toward TA. Although NtSGT expression is known to be induced by biotrophic stress, it was also induced by wounding stress in the same manner as OsSGT. These results indicate that this glucosyltransferase is important not only in biotrophic stress but also for wounding stress. It was found that this enzyme is dually functional, with activity both toward TA and SA.     

Distinct Metabolic Flow Enables Large-Scale Purification of Mouse and Human Pluripotent Stem Cell-Derived Cardiomyocytes

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Highlights? Integrated “omic” analyses reveal metabolic hallmarks in ESCs and cardiomyocytes ? Glucose depletion and lactate supplementation can purify PSC-derived cardiomyocytes ? Cardiomyocytes can use lactate efficiently for energy metabolism ? Transplanted purified human ESC-derived cardiomyocytes do not form tumors     

Quantifying epistatic interactions among the components constituting the protein translation system

In principle, the accumulation of knowledge regarding the molecular basis of biological systems should allow the development of large‐scale kinetic models of their functions. However, the development of such models requires vast numbers of parameters, which are difficult to obtain in practice. Here, we used an in vitro translation system, consisting of 69 defined components, to quantify the epistatic interactions among changes in component concentrations through Bahadur expansion, thereby obtaining a coarse‐grained model of protein synthesis activity. Analyses of the data measured using various combinations of component concentrations indicated that the contributions of larger than 2‐body inter‐component epistatic interactions are negligible, despite the presence of larger than 2‐body physical interactions. These findings allowed the prediction of protein synthesis activity at various combinations of component concentrations from a small number of samples, the principle of which is applicable to analysis and optimization of other biological systems. Moreover, the average ratio of 2‐ to 1‐body terms was estimated to be as small as 0.1, implying high adaptability and evolvability of the protein translation system.     

Genetic Association of Human Leukocyte Antigens with Chronicity or Resolution of Hepatitis B Infection in Thai Population

Background

Previous studies showed that single nucleotide polymorphisms (SNPs) in the HLA-DP, TCF19 and EHMT2 genes may affect the chronic hepatitis B (CHB). To predict the degree of risk for chronicity of HBV, this study determined associations with these SNPs.

Methods

The participants for this study were defined into 4 groups; HCC (n = 230), CHB (n = 219), resolved HBV infection (n = 113) and HBV uninfected subjects (n = 123). The HLA-DP SNPs (rs3077, rs9277378 and rs3128917), TCF19 SNP (rs1419881) and EHMT2 SNP (rs652888) were genotyped.

Results

Due to similar distribution of genotype frequencies in HCC and CHB, we combined these two groups (HBV carriers). The genotype distribution in HBV carriers relative to those who resolved HBV showed that rs3077 and rs9277378 were significantly associated with protective effects against CHB in minor dominant model (OR = 0.45, p<0.001 and OR = 0.47, p<0.001). The other SNPs rs3128917, rs1419881 and rs652888 were not associated with HBV carriers.

Conclusions

Genetic variations of rs3077 and rs9277378, but not rs3128917, rs1419881 and rs652888, were significantly associated with HBV carriers relative to resolved HBV in Thai population.     

Robust circadian rhythms in organoid cultures from PERIOD2::LUCIFERASE mouse small intestine

Disruption of circadian rhythms is a risk factor for several human gastrointestinal (GI) diseases, ranging from diarrhea to ulcers to cancer. Four-dimensional tissue culture models that faithfully mimic the circadian clock of the GI epithelium would provide an invaluable tool to understand circadian regulation of GI health and disease. We hypothesized that rhythmicity of a key circadian component, PERIOD2 (PER2), would diminish along a continuum from ex vivo intestinal organoids (epithelial ‘miniguts’), nontransformed mouse small intestinal epithelial (MSIE) cells and transformed human colorectal adenocarcinoma (Caco-2) cells. Here, we show that bioluminescent jejunal explants from PERIOD2::LUCIFERASE (PER2::LUC) mice displayed robust circadian rhythms for >72 hours post-excision. Circadian rhythms in primary or passaged PER2::LUC jejunal organoids were similarly robust; they also synchronized upon serum shock and persisted beyond 2 weeks in culture. Remarkably, unshocked organoids autonomously synchronized rhythms within 12 hours of recording. The onset of this autonomous synchronization was slowed by >2 hours in the presence of the glucocorticoid receptor antagonist RU486 (20 μM). Doubling standard concentrations of the organoid growth factors EGF, Noggin and R-spondin enhanced PER2 oscillations, whereas subtraction of these factors individually at 24 hours following serum shock produced no detectable effects on PER2 oscillations. Growth factor pulses induced modest phase delays in unshocked, but not serum-shocked, organoids. Circadian oscillations of PER2::LUC bioluminescence aligned with Per2 mRNA expression upon analysis using quantitative PCR. Concordant findings of robust circadian rhythms in bioluminescent jejunal explants and organoids provide further evidence for a peripheral clock that is intrinsic to the intestinal epithelium. The rhythmic and organotypic features of organoids should offer unprecedented advantages as a resource for elucidating the role of circadian rhythms in GI stem cell dynamics, epithelial homeostasis and disease.KEY WORDS: Circadian rhythm, Intestinal organoid, PERIOD2, R-spondin, RU486     

Detection of Genome Donor Species of Neglected Tetraploid Crop Vigna reflexo-pilosa (Créole Bean), and Genetic Structure of Diploid Species Based on Newly Developed EST-SSR Markers from Azuki Bean (Vigna angularis)

Vigna reflexo-pilosa, which includes a neglected crop, is the only one tetraploid species in genus Vigna. The ancestral species that make up this allotetraploid species have not conclusively been identified, although previous studies suggested that a donor genome of V. reflexo-pilosa is V. trinervia. In this study, 1,429 azuki bean EST-SSR markers were developed of which 38 EST-SSR primer pairs that amplified one product in diploid species and two discrete products in tetraploid species were selected to analyze 268 accessions from eight taxa of seven Asian Vigna species including V. reflexo-pilosa var. glabra, V. reflexo-pilosa var. reflexo-pilosa, V. exilis, V. hirtella, V. minima, V. radiata var. sublobata, V. tenuicaulis and V. trinervia to identify genome donor of V. reflexo-pilosa. Since both diploid and tetraploid species were analyzed and each SSR primer pair detected two loci in the tetraploid species, we separated genomes of the tetraploid species into two different diploid types, viz. A and B. In total, 445 alleles were detected by 38 EST-SSR markers. The highest gene diversity was observed in V. hirtella. By assigning the discrete PCR products of V. reflexo-pilosa into two distinguished genomes, we were able to identify the two genome donor parents of créole bean. Phylogenetic and principal coordinate analyses suggested that V. hirtella is a species complex and may be composed of at least three distinct taxa. Both analyses also clearly demonstrated that V. trinervia and one taxon of V. hirtella are the genome donors of V. reflexo-pilosa. Gene diversity indicates that the evolution rate of EST-SSRs on genome B of créole bean might be faster than that on genome A. Species relationship among the Vigna species in relation to genetic data, morphology and geographical distribution are presented.     

Diabetes enhances dental caries and apical periodontitis in caries-susceptible WBN/KobSlc rats

Many epidemiologic studies have suggested that diabetes may be an important risk factor for periodontal disease. To determine whether diabetes induces or enhances periodontal disease or dental caries, dental tissue from diabetic male and nondiabetic female WBN/KobSlc rats and male and female age-matched nondiabetic F344 rats was analyzed morphologically and morphometrically for these 2 types of lesions. Soft X-ray examination revealed that the incidence and severity of both molar caries and alveolar bone resorption were much higher in male WBN/KobSlc rats with chronic diabetes than in nondiabetic female rats of the same strain. Histopathologic examination showed that dental caries progressed from acute to subacute inflammation due to bacterial infections and necrosis in the pulp when the caries penetrated the dentin. In the most advanced stage of dental caries, inflammatory changes caused root abscess and subsequent apical periodontitis, with the formation of granulation tissue around the dental root. Inflammatory changes resulted in resorption of alveolar bone and correlated well with the severity of molar caries. Our results suggest that diabetic conditions enhance dental caries in WBN/KobSlc rats and that periodontal lesions may result from the apical periodontitis that is secondary to dental caries.     

Genomic profiling of submucosal-invasive gastric cancer by array-based comparative genomic hybridization

Genomic copy number aberrations (CNAs) in gastric cancer have already been extensively characterized by array comparative genomic hybridization (array CGH) analysis. However, involvement of genomic CNAs in the process of submucosal invasion and lymph node metastasis in early gastric cancer is still poorly understood. In this study, to address this issue, we collected a total of 59 tumor samples from 27 patients with submucosal-invasive gastric cancers (SMGC), analyzed their genomic profiles by array CGH, and compared them between paired samples of mucosal (MU) and submucosal (SM) invasion (23 pairs), and SM invasion and lymph node (LN) metastasis (9 pairs). Initially, we hypothesized that acquisition of specific CNA(s) is important for these processes. However, we observed no significant difference in the number of genomic CNAs between paired MU and SM, and between paired SM and LN. Furthermore, we were unable to find any CNAs specifically associated with SM invasion or LN metastasis. Among the 23 cases analyzed, 15 had some similar pattern of genomic profiling between SM and MU. Interestingly, 13 of the 15 cases also showed some differences in genomic profiles. These results suggest that the majority of SMGCs are composed of heterogeneous subpopulations derived from the same clonal origin. Comparison of genomic CNAs between SMGCs with and without LN metastasis revealed that gain of 11q13, 11q14, 11q22, 14q32 and amplification of 17q21 were more frequent in metastatic SMGCs, suggesting that these CNAs are related to LN metastasis of early gastric cancer. In conclusion, our data suggest that generation of genetically distinct subclones, rather than acquisition of specific CNA at MU, is integral to the process of submucosal invasion, and that subclones that acquire gain of 11q13, 11q14, 11q22, 14q32 or amplification of 17q21 are likely to become metastatic.     

Elimination of hepatitis C virus from hepatocytes by a selective activation of therapeutic molecules

To eliminate hepatitis C virus (HCV) from infected hepatocytes, we generated two therapeutic molecules specifically activated in cells infected with HCV. A dominant active mutant of interferon (IFN) regulatory factor 7 (IRF7) and a negative regulator of HCV replication, VAP-C (Vesicle-associated membrane protein-associated protein subtype C), were fused with the C-terminal region of IPS-1 (IFNβ promoter stimulator-1), which includes an HCV protease cleavage site that was modified to be localized on the ER membrane, and designated cIRF7 and cVAP-C, respectively. In cells expressing the HCV protease, cIRF7 was cleaved and the processed fragment was migrated into the nucleus, where it activated various IFN promoters, including promoters of IFNα6, IFNβ, and IFN stimulated response element. Activation of the IFN promoters and suppression of viral RNA replication were observed in the HCV replicon cells and in cells infected with the JFH1 strain of HCV (HCVcc) by expression of cIRF7. Suppression of viral RNA replication was observed even in the IFN-resistant replicon cells by the expression of cIRF7. Expression of the cVAP-C also resulted in suppression of HCV replication in both the replicon and HCVcc infected cells. These results suggest that delivery of the therapeutic molecules into the liver of hepatitis C patients, followed by selective activation of the molecules in HCV-infected hepatocytes, is a feasible method for eliminating HCV.