ABSTRACT
DNA methylation is an important epigenetic regulator of gene expression. Application of 5-azacytidine (a methylation inhibitor) significantly promoted bud sprouting rate and the elongation of branches and leaves in "Luhehong" and "Fengdanbai." In total, 11,166 and 11,443 fragments were obtained by methylation-sensitive amplified polymorphism (MSAP) analysis during chilling-induced dormancy release in the two varieties, respectively. Total methylation levels were high in dormant buds, mainly for hemimethylation, which were slowly increased by short-term chilling (7 days) and decreased by long-term chilling. Compared with 0 day, the ratio of the methylation downregulated group increased during dormancy release, whereas that of the upregulated group declined gradually. These variations were consistent with the dynamic expressions of DNA methyltransferase/demethylase genes and their enzyme activity changes. In total, 13 polymorphic MSAP fragments were similar to known proteins (E-value <1e-5), and their methylation statuses were consistent with their expression patterns. The expression change of PsCWH, encoding cell wall hydrolase, might be due to DNA methylation ratios of CpG sites identified by bisulfite sequencing. These results indicated that chilling accumulation promoted bud dormancy release and sprouting through DNA methylation modification of specific genes. This study would provide new insights into the molecular mechanism underlying dormancy release in tree peony.
Subject(s)
Epigenesis, Genetic/genetics , Paeonia/genetics , Plant Dormancy/genetics , Azacitidine/pharmacology , China , Computational Biology/methods , DNA/genetics , DNA Methylation/genetics , Epigenomics/methods , Flowers/genetics , Gene Expression Regulation, Plant/genetics , Paeonia/metabolism , Plant Dormancy/physiology , Trees/genetics , Trees/metabolismABSTRACT
BACKGROUND: Long non-coding RNAs (lncRNAs) are transcribed pervasively in the genome and act to regulate chromatin remodeling and gene expression. Dysregulated lncRNA expression has been reported in many cancers, but the role of lncRNAs in esophageal cancer (EC) has so far remained poorly understood. In this study, we aimed to understand the effect of lncRNA LINC01234 on EC development through competitively binding to microRNA-193a-5p (miR-193a-5p). METHODS: The Gene Expression Omnibus (GEO) database was used for microarray-based EC expression profiling. Gain- and loss-of-function analyses were carried out in human EC-derived Eca-109 and EC9706 cells. Expression analyses of miR-193a-5p, LINC01234, CCNE1, caspase-3, p21, Bax, cyclinD1 and Bcl-2 were performed using RT-qPCR and Western blotting. Cell proliferation, colony formation and apoptosis analyses were carried out using MTT, Hoechst 33258 and flow cytometry assays. A xenograft EC model in nude mice was used to evaluate in vivo tumor growth and CCNE1 expression. RESULTS: Microarray-based analyses revealed that LINC01234 expression was increased in primary EC samples, whereas that of miR-193a-5p was decreased. We found that CCNE1 was a target of miR-193a-5p and that LINC01234, in turn, sponges miR-193a-5p. After treatment with si-LINC01234 or miR-193a-5p mimic, EC cells (Eca-109 and EC9706) exhibited cyclinD1 and Bcl-2 downregulation, and caspase-3, p21, Bax and cleaved caspase-3 upregulation. LINC01234 silencing or miR-193a-5p upregulation resulted in decreased proliferation and colony formation, and increased apoptosis of EC cells. In addition, LINC01234 silencing or miR-193a-5p upregulation resulted in reduced in vivo EC tumor growth and CCNE1 expression in nude mice. CONCLUSIONS: We found that silencing of LINC01234 suppresses EC development by inhibiting CCNE1 through competitively binding to miR-193a-5p, which suggests that LINC01234 may represent a novel target for EC therapy.
Subject(s)
Cyclin E/metabolism , Down-Regulation/genetics , Esophageal Neoplasms/genetics , Gene Expression Regulation, Neoplastic , Gene Silencing , MicroRNAs/metabolism , Oncogene Proteins/metabolism , RNA, Long Noncoding/metabolism , Animals , Apoptosis/genetics , Cell Line, Tumor , Cell Proliferation/genetics , Esophageal Neoplasms/pathology , Humans , Mice, Nude , MicroRNAs/genetics , Models, Biological , RNA, Long Noncoding/genetics , Up-Regulation/genetics , Xenograft Model Antitumor AssaysABSTRACT
Bud endodormancy in tree peony is a growth cessation-like state, and sufficient chilling perception is necessary to break it. In this study, 120 plants were subjected to 0-4⯰C climate chamber for 0-28â¯d with a weekly interval, morphology and structure changes of buds were studied with a scanning electron microscope (SEM) and paraffin sections during the dormancy process. Dormancy status was evaluated after being transferred to greenhouse for 30â¯d. Results showed that the diameter of the buds gradually expanded, along with continuous elongation of sepals, petals, stamens and carpels in the chilling accumulation process. Notably, dormancy release was marked with the establishment of xylem vessels in lateral vein of the petal. Meanwhile, DNA methylation was detected by HPLC and immunochemical technology, aimed to illuminate the role of DNA methylation in the dormancy release, we found that 5â¯mC level fell from 39.4% to 24.2% after exposed to 28â¯d chilling. These results were consistent with the immunochemical analysis, and inversely related to the sprouting rate after being moved to greenhouse for 30â¯d. Exogenous application of 5azaC (5-azacytidine) decreased DNA methylation level, accompanied by an improved bud sprouting capacity, while the effect of SAM (S-adenosylmethionine) was the opposite. In summary, prolonged chilling was accompanied by further differentiation and development of the compound bud, which resulted in DNA hypomethylation and promoted dormancy release in tree peony.
