The epitranscriptomic mark N⁶-methyladenosine (m⁶A), which is the predominant internal modification in RNA, is important for plant responses to diverse stresses. Multiple environmental stresses caused by the tea-withering process can greatly influence the accumulation of specialized metabolites and the formation of tea flavor. However, the effects of the m⁶A-mediated regulatory mechanism on flavor-related metabolic pathways in tea leaves remain relatively uncharacterized. We performed an integrated RNA methylome and transcriptome analysis to explore the m⁶A-mediated regulatory mechanism and its effects on flavonoid and terpenoid metabolism in tea (Camellia sinensis) leaves under solar-withering conditions. Dynamic changes in global m⁶A level in tea leaves were mainly controlled by two m⁶A erasers (CsALKBH4A and CsALKBH4B) during solar-withering treatments. Differentially methylated peak-associated genes following solar-withering treatments with different shading rates were assigned to terpenoid biosynthesis and spliceosome pathways. Further analyses indicated that CsALKBH4-driven RNA demethylation can directly affect the accumulation of volatile terpenoids by mediating the stability and abundance of terpenoid biosynthesis-related transcripts and also indirectly influence the flavonoid, catechin, and theaflavin contents by triggering alternative splicing-mediated regulation. Our findings revealed a novel layer of epitranscriptomic gene regulation in tea flavor-related metabolic pathways and established a link between the m⁶A-mediated regulatory mechanism and the formation of tea flavor under solar-withering conditions.
RNA/metabolism* ; Epigenome ; Plant Proteins/metabolism* ; Plant Leaves/metabolism* ; Camellia sinensis/metabolism* ; Flavonoids ; Terpenes/metabolism* ; Tea/metabolism* ; Gene Expression Regulation, Plant

Oolong tea is a semi-fermented tea with strong flavor, which is widely favored by consumers because of its floral and fruity aroma as well as fresh and mellow taste. During the processing of oolong tea, withering is the first indispensable process for improving flavor formation. However, the molecular mechanism that affects the flavor formation of oolong tea during withering remains unclear. Transcriptome sequencing was used to analyze the difference among the fresh leaves, indoor-withered leaves and solar-withered leaves of oolong tea. A total of 10 793 differentially expressed genes were identified from the three samples. KEGG enrichment analysis showed that the differentially expressed genes were mainly involved in flavonoid synthesis, terpenoid synthesis, plant hormone signal transduction and spliceosome pathways. Subsequently, twelve differentially expressed genes and four differential splicing genes were identified from the four enrichment pathways for fluorescence quantitative PCR analysis. The results showed that the expression patterns of the selected genes during withering were consistent with the results in the transcriptome datasets. Further analysis revealed that the transcriptional inhibition of flavonoid biosynthesis-related genes, the transcriptional enhancement of terpenoid biosynthesis-related genes, as well as the jasmonic acid signal transduction and the alternative splicing mechanism jointly contributed to the flavor formation of high floral and fruity aroma and low bitterness in solar-withered leaves. The results may facilitate better understanding the molecular mechanisms of solar-withering treatment in flavor formation of oolong tea.
Camellia sinensis/genetics* ; Gene Expression Profiling ; Plant Leaves ; Plant Proteins/metabolism* ; Taste ; Tea ; Transcriptome/genetics*

Objective To study the relationship between single subcortical small infarction (SSSI) in middle cerebral artery (MCA) and large artery atheromatous sclerosis (LAAS).Methods One hundred and sixty patients with acute SSSI in MCA admitted to our hospital from January 2014 to December 2015 were divided into proximal infarction group (n=98) and distal infarction group (n=62).Their demographic,clinical,imaging data and National Institutes of Health Stroke Scale (NIHSS) score were recorded on admision.Results The incidence of coronary heart disease,NIHSS score,and contralateral intracranial artery stenosis were significantly higher while the incidence of proximal infarction,old lacunar infarction,and periventricular leukoaraiosis was significantly lower in proximal infarction group than in distal infarction group (21.4％ vs 3.2％,P=0.001;3 vs 2,P=0.030;19.4％ vs 1.6％,P=0.002;10.2％ vs 29.0％,P=0.002;16.3％ vs 33.9％,P=0.010).Logistic regression analysis showed that proximal infarction was independently related with old lacunar infarction,contralateral intracranial artery stenosis and periventricular leukoaraiosis (OR=0.270,95％CI:0.100-0.726,P=0.009;OR=4.500,95％CI:1.541-15.012,P=0.018;OR=0.325,95％CI:0.127-0.834,P=0.019).Conclusion Distal SSSI in MCA is related with small vessel disease while proximal SSSI in MCA is related with LAAS.

Objective·To investigate the role of collagen triple helix repeat containing-1 (CTHRC1) in ovarian cancer cell metastasis and the related mechanism. Methods·The expression of CTHRC1 in ovarian cancer cells was detected by Western blotting. The cell line which had high expression of CTHRC1 was transfected with a CTHRC1 specific shRNA, and the lenti-CTHRC1 was used to overexpress CTHRC1 in the cell line whose expression of CTHRC1 was very low. Then the expression of Slug and MMP-2 was assessed. Transwell assay was used to determine the migration and invasion capability of ovarian cancer cells after transfection. Results·The expression of CTHRC1 in HO8910 cells was the lowest, while the CTHRC1 protein level was dramatically increased after transfection of lenti-CTHRC1. Meanwhile, there was a distinct rise of the migration and invasion ability, as well as the expression of Slug and MMP-2 (all P<0.05). Conversely, CaOV3 cells had a higher protein expression of CTHRC1. By using lenti-shCTHRC1, a remarkable knockdown of CTHRC1 was obtained. Likewise, the capability of migration and invasion was decreased, and the Slug and MMP-2 expression was reduced (all P<0.05). Conclusion·CTHRC1 might positively regulate Slug and MMP-2 to promote ovarian cancer cell metastasis.