Investigating Mitotic Inheritance of Histone Modifications Using Tethering Strategies.
Investigating Mitotic Inheritance of Histone Modifications Using Tethering Strategies.
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DOI：
10.1007/978-1-0716-2481-4_18
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发表时间：
2022
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组蛋白的共价和可逆修饰使细胞能够在不改变其遗传蓝图的情况下建立可遗传的基因表达模式。表观遗传机制以两种不同的方式调节基因表达：（1）建立，这取决于序列特异性DNA或RNA结合蛋白，其将组蛋白修饰酶募集到独特的基因组位点，和（2）维持，这是序列无关的，并且取决于DNA复制期间预先存在的染色质状态的自主繁殖。在基因组中，只有大量组蛋白修饰的子集是可遗传的。在这里，我们描述了一种合成生物学的方法来拴系组蛋白修饰酶工程染色质状态的活细胞，并评估其潜在的有丝分裂遗传。In S.粟酒裂殖酵母，融合的H3K9甲基转移酶，Clr 4，四环素诱导TetR DNA结合结构域促进快速和可逆的控制异染色质装配。我们描述了一个框架，成功地实现了诱导异染色质建立系统，并评估其分子特性。我们预计，我们创新的遗传策略将广泛适用于发现蛋白质复合物和异染色质相关因子的功能分离等位基因，这些因子在表观遗传中具有独特的作用。
The covalent and reversible modification of histones enables cells to establish heritable gene expression patterns without altering their genetic blueprint. Epigenetic mechanisms regulate gene expression in two separate ways: (1) establishment, which depends on sequence-specific DNA- or RNA-binding proteins that recruit histone-modifying enzymes to unique genomic loci, and (2) maintenance, which is sequence-independent and depends on the autonomous propagation of preexisting chromatin states during DNA replication. Only a subset of the vast repertoire of histone modifications in the genome is heritable. Here, we describe a synthetic biology approach to tether histone-modifying enzymes to engineer chromatin states in living cells and evaluate their potential for mitotic inheritance. In S. pombe, fusing the H3K9 methyltransferase, Clr4, to the tetracycline-inducible TetR DNA-binding domain facilitates rapid and reversible control of heterochromatin assembly. We describe a framework to successfully implement an inducible heterochromatin establishment system and evaluate its molecular properties. We anticipate that our innovative genetic strategy will be broadly applicable to the discovery of protein complexes and separation-of-function alleles of heterochromatin-associated factors with unique roles in epigenetic inheritance.