ROLE OF CYTOSINE METHYLATION IN C. CINEREUS

灰盖鬼伞胞嘧啶甲基化的研究 已知CpG甲基化参与基因沉默， 负责基因组印记和X染色体失活。 这 这项工作解决了真菌中胞嘧啶甲基化的后果， 灰盖鬼伞系统，目前正在研究其位置和 色氨酸合成酶中5-甲基胞嘧啶（m5 C）残基的范围 基因（trpi）。 基因组测序方案已经过完善， 在我们的真菌系统中甲基化。 在这一批判性的反应中， 方案，胞嘧啶在用钠处理后脱氨基为尿嘧啶 亚硫酸氢盐，而m5 C保持不变。 因此，m5 C的存在 残基由序列数据中剩余的胞嘧啶表示。 对同时具有Trp+和Trp-后代的六个四分体的分析使我能够 直接比较基因沉默的 和不沉默的基因 所有DNA的序列分析证明， 胞嘧啶甲基化主要发生在CpG二核苷酸处，如 哺乳动物系统，Trp-DNA具有更大量的甲基化 胞嘧啶与Trp+ DNA相比。 进一步分析将确定 胞嘧啶甲基化具有位点特异性， 甲基化胞嘧啶是必需的，足以使TRP 1 基因失活。 C. cinereus已经被证明是研究 甲基化，这可能会给我们一些线索，关于基因沉默是如何发生在 这一物种和整个真核生物都可能存在。 这可能导致 更好地理解m5 C在控制基因表达中的作用 并进一步阐明其在癌症等疾病中的作用 脆性X综合征

Examining the Role of Cytosine Methylation in Coprinus cinereus CpG methylation is known to be involved in gene silencing and to be responsible for genomic imprinting and X-chromosome inactivation. This work addresses the consequences of cytosine methylation in the fungal system Coprinus cinereus and currently is studying the location and extent of 5-methyl cytosine (m5C) residues in the tryptophan synthetase gene (trpi). The genomic sequencing protocol has been refined to study methylation in our fungal system. In the critical reaction of this protocol, cytosine is deaminated to uracil upon treatment with sodium bisulfite while m5C remains unchanged. Thus, the presence of m5C residues is indicated by the remaining cytosines in the sequence data. Analysis of six tetrads with both Trp+ and Trp- progeny allowed me to make a direct comparison of differences in methylation between silenced and unsilenced genes. Sequence analysis of all DNAs proved that cytosine methylation occurs mostly at the CpG dinucleotide as in mammalian systems, and that Trp- DNA has a larger amount of methylated cytosines as compared to Trp+ DNA. Further analysis will determine if there is site specificity for cytosine methylation and if specific methylated cytosines are necessary and sufficient to render the trp1 gene inactive. C. cinereus has proven to be a model system for studying methylation and may give us clues about how gene silencing occurs in this species and possibly in eukaryotes as a whole. This may lead to a better understanding of the role of m5C in controlling gene expression and further elucidate its role in diseases such as cancer and the Fragile-X syndrome.