Regulation of chromatin topology

染色质是包裹基因组的DNA和蛋白质的基本混合物。细胞中存在几种不同形式的染色化DNA，这些状态决定了哪些DNA序列被读取，哪些被沉默。这可能是细胞中最关键的决定：随之而来的基因表达谱（哪些基因是“开”的，哪些基因是“关”的）是细胞身份和行为的关键决定因素。因此，了解染色质状态如何建立和调节与广泛的科学领域相关，包括细胞生长调节、细胞和生物体发育以及环境-疾病关系的研究。******球状核小体是染色质的基本重复单位；它由8种被称为组蛋白的包装蛋白组成，这些蛋白质被包裹在1.5圈DNA中。在“开启”的基因中，核小体以松散而开放的“串珠”组合形式排列。在“关闭”的基因中，核小体可以组装成更高级的有序结构，称为染色质拓扑结构。这些状态是由一组组蛋白修饰酶动态调节的，这些酶是目前研究热点。***我们发现了一组新的组蛋白修饰酶，称为组蛋白脯氨酸异构酶。这些酶改变核小体“头”的一种组蛋白的形状。在上一个资助期，我们使用体外生化技术和分子遗传学方法证明了组蛋白脯氨酸异构酶在试管中致密染色质并沉默细胞中的特定DNA序列。我们下一个资助期的目标是：1)确定这些致密纤维中的拓扑特性（即核小体的排列），2)了解这种形状变化如何影响关键DNA读取蛋白在活细胞中参与染色质的能力。我们的实验室将是第一个结合尖端合成化学、分子生物学和高功率电子显微镜来构建和分析这些染色质拓扑结构的实验室。然后，工程合成拓扑结构将被用作分子诱饵，以捕获和识别细胞中在这种核小体排列上组装的蛋白质。******从我们的研究中获得的知识将进一步了解所有真核生物基因组是如何控制的。这直接影响到广泛的科学领域，包括细胞生长调节、细胞和生物体发育以及环境-疾病关系的研究。最后，采用了几种先进的集成技术。这将为众多年轻的加拿大理科硕士、博士和本科科学家提供世界级的培训。＊＊＊＊＊＊＊＊

Chromatin is an essential mixture of DNA and protein that packages the genome. Several different forms of chromatinized DNA exist in cells, and these states determine which DNA sequences are read and which are silenced. This is perhaps the most critical decision to be made in a cell: the consequent gene expression profile (which genes are `ON' and which genes are `OFF') is the key determinant of cellular identity and behavior. Learning how chromatin states are established and regulated is therefore relevant to broad scientific fields including the studies of the regulation of cell growth, cell and organism development and environment-disease relationships. ******The globular nucleosome is the fundamental repeating unit of chromatin; it is comprised of eight packaging proteins called histones that are wrapped in 1.5 turns of DNA. In genes that are `ON', nucleosomes are spaced in a loose and open `beads-on-a-string' assembly. In genes that are `OFF' nucleosomes can assemble into higher ordered structures, called chromatin topologies. These states are dynamically regulated by a collection of histone-modifying enzymes that are currently the focus of an intense area of research.***We have identified a new group of histone-modifying enzymes called histone prolyl isomerases. These enzymes alter the shape of one histone protein of the nucleosome `bead'. In the last funding period we used biochemical in vitro techniques and molecular genetic approaches to show that histone prolyl isomerases compact chromatin in a test tube and silence specific DNA sequences in cells. Our goals in the next funding period are: 1) to define the topological properties (ie arrangement of nucleosomes) in these compacted fibers and, 2) to learn how this shape change influences the ability of critical DNA reading proteins to engage chromatin in live cells. Our lab will be the first to combine cutting-edge synthetic chemistry, molecular biology and high-powered electron microscopy to build and analyze these chromatin topologies. Engineered synthetic topologies will then be used as molecular bait to capture and identify proteins that assemble on such nucleosomal arrangements in cells.******The knowledge acquired from our research will further the understanding of how all eukaryotic genomes are controlled. This has direct implications to broad scientific fields including the studies of the regulation of cell growth, cell and organism development and environment-disease relationships. Finally, several highly-advanced and integrated technologies are applied. This will provide world-class training to numerous young Canadian MSc, PhD and undergraduate scientists. ********