Hypermethylation in the promoter regions of flavonoid pathway genes is associated with skin color fading during 'Daihong' apple fruit development.

Apple fruit skin color fading is not well understood although the molecular mechanism of skin color formation is well known. The red-fleshed apple cultivar 'Daihong' (DH) exhibited fading skin color during fruit development despite having a heterozygous R6 allele but lacking Red-TE for red fruit skin. In this study, transcriptomic analysis revealed the expression level of MdMYB10 increased with fruit development whereas reduced expression levels of MdMYBPA1, MdCHS, MdANS, MdUFGT, MdLAR, and MdANR were observed, consistent with decreased levels of chalcone, anthocyanin, catechin, epicatechin, and procyanidin B2. Whole-genome bisulfite sequencing (WGBS) indicated a global gain in cytosine methylation levels and increased methylation in 5' and 3' flanking regions of genes and transposable elements (TEs), and in TE bodies in all CG, CHG and CHH contexts, especially the mCHH context, during fruit development. The increased DNA methylation was attributed to reduced expression levels of DNA demethylase genes, including MdDME1, MdROS1, and MdROS2. Association analysis revealed a significant negative correlation between promoter methylation levels of MdCHS, MdCHI, MdMYBPA1, and their respective transcript levels, as well as a negative correlation between promoter methylation levels of MdCHS, MdCHI, MdANR, and MdFLS, and the content of chalcones, naringenin-7-glucoside, epicatechin, and quercetin. Treatment with the DNA demethylation agent 5-aza-2'-deoxycytidine verified the negative correlation between DNA methylation and gene expression within the flavonoid pathway. These findings suggest that hypermethylation in promoter regions of genes of the flavonoid biosynthesis pathway is associated with the reduction of gene expression and flavonoid content, and fruit skin color fading during DH apple development.

Relationship Between Serum Indirect Bilirubin Levels and Cardiovascular Events and All-Cause Mortality in Maintenance Hemodialysis Patients.

Purpose: Unconjugated bilirubin is one of the most endogenous antioxidant substances. Mildly elevated total bilirubin concentrations may protect against cardiovascular disease and total death. However, most studies only focused on the association between serum total bilirubin and the risk of cardiovascular disease and total death. This study aimed to investigate the relationship between serum indirect bilirubin (IBIL) and the cardiovascular events in maintenance hemodialysis patients. Patients and Methods : This retrospective cohort study included 284 maintenance hemodialysis patients. Patients were divided into two groups according to the median IBIL level: high IBIL group (IBIL ≥3.0 µmol/L) and low IBIL group (IBIL <3.0 µmol/L). All demographic and laboratory data were recorded at baseline. The endpoint was cardiovascular events and all-cause mortality. Results: During the median follow-up time of 62 months, 96 patients developed cardiovascular disease. There were 134 deaths. In Kaplan-Meier analysis curves, the risk of cardiovascular events in the low IBIL group was significantly higher than high IBIL group (P < 0.001). In multivariate Cox regression analysis, the risk of cardiovascular events in high IBIL group was 0.484 times (95% CI 0.278-0.844, P = 0.010) the risk in low IBIL group. However, there was no significant association between serum IBIL level and all-cause mortality (P = 0.269). Conclusion: Our findings suggest that lower circulating IBIL levels were associated with the increased risk of cardiovascular events in maintenance hemodialysis patients.

Transcriptome and methylome changes in two contrasting mungbean genotypes in response to drought stress.

BACKGROUND: Due to drought stress, the growth, distribution, and production of mungbean is severely restricted. Previous study combining physiological and transcriptomic data indicated different genotypes of mungbean exhibited variable responses when exposed to drought stress. Aside from the genetic variation, the modifications of environmentally induced epigenetics alterations on mungbean drought-stress responses were still elusive. RESULTS: In this study, firstly, we compared the drought tolerance capacity at seedling stage by detecting physiological parameters in two contrasting genotypes wild mungbean 61 and cultivar 70 in response to drought stress. We found that wild mungbean 61 showed lower level of MDA and higher levels of POD and CAT, suggesting wild mungbean 61 exhibited stronger drought resistance. Transcriptomic analysis indicated totally 2859 differentially expressed genes (DEGs) were detected when 70 compared with 61 (C70 vs C61), and the number increased to 3121 in the comparison of drought-treated 70 compared with drought-treated 61 (D70 vs D61). In addition, when drought-treated 61 and 70 were compared with their controls, the DEGs were 1117 and 185 respectively, with more down-regulated DEGs than up-regulated in D61 vs C61, which was opposite in D70 vs C70. Interestingly, corresponding to this, after drought stress, more hypermethylated differentially methylated regions (DMRs) in 61 were detected and more hypomethylated DMRs in 70 were detected. Further analysis suggested that the main variations between 61 and 70 existed in CHH methylation in promoter. Moreover, the preference of methylation status alterations in D61 vs C61 and D70 vs C70 also fell in CHH sequence context. Further analysis of the correlation between DMRs and DEGs indicated in both D61 vs C61 and D70 vs C70, the DMRs in gene body was significantly negatively correlated with DEGs. CONCLUSIONS: The physiological parameters in this research suggested that wild mungbean 61 was more resistant to drought stress, with more hypermethylated DMRs and less hypomethylated DMRs after drought stress, corresponding to more down-regulated DEGs than up-regulated DEGs. Among the three DNA methylation contexts CG, CHG, and CHH, asymmetric CHH contexts were more dynamic and prone to be altered by drought stress and genotypic variations.

Effect of Nanocrystallization of Anthocyanins Extracted from Two Types of Red-Fleshed Apple Varieties on Its Stability and Antioxidant Activity.

Red-fleshed apple (Malus sieversii f. neidzwetzkyana (Dieck) Langenf) has attracted more and more attention due to its enriched anthocyanins and high antioxidant activity. In this study we extracted total anthocyanins and phenols from two types of red-fleshed apples-Xinjing No.4 (XJ4) and Red Laiyang (RL)-to study the stability and antioxidant activity of anthocyanins after encapsulation onto Corn Starch Nanoparticles (CSNPs). The results indicated the anthocyanins and total phenol levels of XJ4 were 2.96 and 2.25 times higher than those of RL respectively. The anthocyanin concentration and loading time had a significant effect on CSNPs encapsulation, and XJ4 anthocyanins always showed significantly higher loading capacity than RL. After encapsulation, the morphology of RL-CSNPs and XJ4-CSNPs was still spherical with a smooth surface as CSNPs, but the particle size increased compared to CSNPs especially for RL-CSNPs. Different stress treatments including UV light, pH, temperature, and salinity suggested that XJ4-CSNPs exhibited consistently higher stability than RL-CSNPs. A significantly enhanced free radical scavenging rate under stress conditions was observed, and XJ4-CSNPs had stronger antioxidant activity than RL-CSNPs. Furthermore, XJ4-CSNPs exhibited a slower released rate than RL-CSNPs in simulated gastric (pH 2.0) and intestinal (pH 7.0) environments. Our research suggests that nanocrystallization of anthocyanins is an effective method to keep the anthocyanin ingredients intact and active while maintaining a slow release rate. Compared to RL, encapsulation of XJ4 anthocyanins has more advantages, which might be caused by the significant differences in the metabolites of XJ4. These findings give an insight into understanding the role of nanocrystallization using CSNPs in enhancing the antioxidant ability of anthocyanins from different types of red-fleshed apples, and provide theoretical foundations for red-fleshed apple anthocyanin application.