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

CC chemokine receptor 4 (CCR4) is a kind of G-protein-coupled receptor, which plays a pivotal role in allergic inflammation. The interaction between 2-(2-(4-chloro-phenyl)-5-{[(naphthalen-1-ylmethyl)-carbamoyl]-methyl}-4-oxo-thiazolidin-3-yl)-N-(3-morpholin-4-yl-propyl)-acetamide (S009) and the N-terminal extracellular tail (ML40) of CCR4 has been validated to be high affinity by capillary zone electrophoresis (CZE). The S009 is a known CCR4 antagonist. Now, a series of new thiourea derivatives have been synthesized. Compared with positive control S009, they were screened using ML40 as target by CZE to find some new drugs for allergic inflammation diseases. The synthesized compounds XJH-5, XJH-4, XJH-17 and XJH-1 displayed the interaction with ML40, but XJH-9, XJH-10, XJH-11, XJH-12, XJH-13, XJH-14, XJH-3, XJH-8, XJH-6, XJH-7, XJH-15, XJH-16 and XJH-2 did not bind to ML40.Both qualification and quantification characterizations of the binding were determined. The affinity of the four compounds was valued by the binding constant, which was similar with the results of chemotactic experiments. The established CEZ method is capable of sensitive and fast screening for a series of lactam analogs in the drug discovery for allergic inflammation diseases.