Regulation of histone methylation

在真核生物中，DNA被包装成染色质，一种由DNA、组蛋白和非组蛋白组成的核蛋白结构。 染色质对于基因表达的保真度是重要的，并且其结构可以通过向组蛋白添加翻译后修饰（PTM）来调节。 一种这样的修饰，组蛋白H3在赖氨酸36处的甲基化，通过RNA聚合酶II的传递靶向活性基因，并且这种PTM可以说是所有真核生物中生产性转录的最公认的标志。 然而，尽管对H3K36甲基化的建立和功能背后的机制进行了十多年的研究，但令人惊讶的是，人们对这种修饰在基因抑制后如何被去除知之甚少。 本申请中提出的研究将揭示在基因阻遏期间参与去除H3K36甲基化的酶和途径。 我们将使用S。由于参与酵母和其它真核生物之间H3K36甲基化调节的基因的极好的保守性，以及该模式生物的遗传易处理性，因此，本发明的目的是将酿酒酵母作为模式生物。 所使用的技术包括ChIP（染色质免疫沉淀），qPCR（定量聚合酶链反应），深度测序，生物信息学，蛋白质纯化，酶动力学和酵母遗传学的文库制备，因此由我的NSERC资助的学生将学习在生命科学研究的多个领域备受追捧的各种技能。

In eukaryotes DNA is packaged into chromatin, a nucleoprotein structure consisting of DNA, histones, and non-histone proteins. Chromatin is important for the fidelity of gene expression and its structure can be regulated by the addition of post-translational modifications (PTMs) to histones. One such modification, methylation of histone H3 at lysine 36, is targeted to active genes by the passage of RNA polymerase II, and this PTM is arguably the most recognized hallmark of productive transcription in all eukaryotes. However, despite over a decade worth of research into the mechanisms behind the establishment and function of H3K36 methylation, there is surprisingly little known about how this modification is removed following gene repression. The research proposed in this application will reveal the enzymes and pathways involved in removal of H3K36 methylation during gene repression. We will use S. cerevisiae as a model organism due to the excellent conservation of genes involved in regulation of H3K36 methylation between yeast and other eukaryotes, and the genetic tractability of this model organism. The techniques used include ChIP (chromatin immunoprecipitation), qPCR (quantitative polymerase chain reaction), library preparation for deep sequencing, bioinformatics, protein purification, enzyme kinetics, and yeast genetics, and thus students supported by my NSERC grant will learn a diverse set of skills that are highly sought after in multiple fields of life science research.