Anthocyanins play an important role in the growth of plants, and are beneficial to human health. In plants, the MYB-bHLH-WD40 (MBW) complex activates the genes for anthocyanin biosynthesis. However, in rice, the WD40 regulators remain to be conclusively identified. Here, a crucial anthocyanin biosynthesis gene was fine mapped to a 43.4-kb genomic region on chromosome 2, and a WD40 gene OsTTG1 (Oryza sativa TRANSPARENT TESTA GLABRA1) was identified as ideal candidate gene. Subsequently, a homozygous mutant (osttg1) generated by CRISPR/Cas9 showed significantly decreased anthocyanin accumulation in various rice organs. OsTTG1 was highly expressed in various rice tissues after germination, and it was affected by light and temperature. OsTTG1 protein was localized to the nucleus, and can physically interact with Kala4, OsC1, OsDFR and Rc. Furthermore, a total of 59 hub transcription factor genes might affect rice anthocyanin biosynthesis, and LOC_Os01g28680 and LOC_Os02g32430 could have functional redundancy with OsTTG1. Phylogenetic analysis indicated that directional selection has driven the evolutionary divergence of the indica and japonica OsTTG1 alleles. Our results suggest that OsTTG1 is a vital regulator of anthocyanin biosynthesis, and an important gene resource for the genetic engineering of anthocyanin biosynthesis in rice and other plants.

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BackgroundActivation of amoeboid microglial cells (AMC) and its related inflammatory response have been linked to the periventricular white matter damage after hypoxia in neonatal brain. Hypoxia increases free ATP in the brain and then induces various effects through ATP receptors. The present study explored the possible mechanism in ATP induced AMC activation in hypoxia.ResultsWe first examined the immunoexpression of P2X4, P2X7 and P2Y12 in the corpus callosum (CC) and subependyma associated with the lateral ventricles where both areas are rich in AMC. Among the three purinergic receptors, P2X4 was most intensely expressed. By double immunofluorescence, P2X4 was specifically localized in AMC (from P0 to P7) but the immunofluorescence in AMC was progressively diminished with advancing age (P14). It was further shown that P2X4 expression was noticeably enhanced in P0 day rats subjected to hypoxia and killed at 4, 24, 72 h and 7 d versus their matching controls by double labeling and western blotting analysis. P2X4 expression was most intense at 7 d whence the inflammatory response was drastic after hypoxia. We then studied the association of P2X4 with cytokine release in AMC after hypoxic exposure. In primary microglial cells exposed to hypoxia, IL-1β and TNF-α protein levels were up-regulated. Blockade of P2X4 receptor with 2', 3'-0-(2, 4, 6-Trinitrophenyl) adenosine 5'-triphosphate, a selective P2X1-7 blocker resulted in partial suppression of IL-1β (24% vs hypoxic group) and TNF-α expression (40% vs hypoxic group). However, pyridoxal phosphate-6-azo (benzene-2, 4-disulfonic acid) tetrasodium salt hydrate, a selective P2X1-3, 5-7 blocker did not exert any significant effect on the cytokine expression.ConclusionsIt is concluded that P2X4 which is constitutively expressed by AMC in postnatal rats was enhanced in hypoxia. Hypoxia induced increase in IL-1β and TNF-α expression was reversed by 2', 3'-0-(2, 4, 6-Trinitrophenyl) adenosine 5'-triphosphate suggesting that P2X4 mediates ATP induced AMC activation and its production of proinflammatory cytokines.

Background and Aims Portopulmonary hypertension (POPH) was previously associated with a single‐nucleotide polymorphism (SNP) rs7175922 in aromatase (cytochrome P450 family 19 subfamily A member 1 [CYP19A1]). We sought to determine whether genetic variants and metabolites in the estrogen signaling pathway are associated with POPH. Approach and Results We performed a multicenter case‐control study. POPH patients had mean pulmonary artery pressure >25 mm Hg, pulmonary vascular resistance >240 dyn‐sec/cm−5, and pulmonary artery wedge pressure ≤15 mm Hg without another cause of pulmonary hypertension. Controls had advanced liver disease, right ventricular (RV) systolic pressure <40 mm Hg, and normal RV function by echocardiography. We genotyped three SNPs in CYP19A1 and CYP1B1 using TaqMan and imputed SNPs in estrogen receptor 1 using genome‐wide markers. Estrogen metabolites were measured in blood and urine samples. There were 37 patients with POPH and 290 controls. Mean age was 57 years, and 36% were female. The risk allele A in rs7175922 (CYP19A1) was significantly associated with higher levels of estradiol (P = 0.02) and an increased risk of POPH (odds ratio [OR], 2.36; 95% confidence interval [CI], 1.12‐4.91; P = 0.02) whereas other SNPs were not. Lower urinary 2‐hydroxyestrogen/16‐α‐hydroxyestrone (OR per 1‐ln decrease = 2.04; 95% CI, 1.16‐3.57; P = 0.01), lower plasma levels of dehydroepiandrosterone‐sulfate (OR per 1‐ln decrease = 2.38; 95% CI, 1.56‐3.85; P < 0.001), and higher plasma levels of 16‐α‐hydroxyestradiol (OR per 1‐ln increase = 2.16; 95% CI, 1.61‐2.98; P < 0.001) were associated with POPH. Conclusions Genetic variation in aromatase and changes in estrogen metabolites were associated with POPH.