The Dosage Compensation Machinery of C. elegans

描述(申请人提供)：凝集素复合体在有丝分裂过程中发挥作用，形成浓缩的有丝分裂染色体，并在间期调节基因的表达。这两个目标都被认为是通过改变染色体的更高顺序结构来实现的。在线虫中，两个不同的凝集素复合体执行这两个功能。有丝分裂凝集素复合体在染色体分离中起作用，而剂量补偿复合体调节两性X染色体上的基因表达，使X连锁基因在两性之间的剂量相等。我们的长期目标是破译蠕虫剂量补偿的分子机制，并确定它与染色体凝聚机制的关系。剂量补偿复合体的生化纯化导致了Cpg-1的发现，Cpg-1是一种新的剂量补偿复合体亚基，它也在染色体分离中发挥作用。为了揭示剂量补偿和染色体分离之间的机制相似性，我们将使用遗传学、生化和显微工具来分析Cpg-1在这两个过程中发挥作用的方式。我们提出的研究的第二部分将分析剂量补偿染色体的染色质纤维。我们发现，与常染色体相比，H_2A的组蛋白变异体HTZ-1在剂量补偿的X染色体上耗尽。我们提出的实验的目标是破译这种耗尽是如何有助于剂量补偿的。为此，我们将研究剂量补偿复合体和HTZ-1与X染色体的关系，无论是在低分辨率(免疫荧光)还是高分辨率(染色质免疫沉淀)。任何生物体的健康发育都有赖于向每个细胞和每个细胞传递正确数量的染色体，然后启动正确的一组基因。将错误数量的染色体传递给细胞，或者不适当地开启或关闭基因，都会导致发育障碍或癌症。染色体的这些功能在很大程度上取决于它们所包含的DNA的结构和组织。更好地了解染色体结构如何决定基因活性水平和确保适当的染色体分离的基本生物学原理，对于理解健康发育和疾病的发展都是重要的。

DESCRIPTION (provided by applicant): The condensin complex functions both during mitosis, to form condensed mitotic chromosomes, and during interphase, to regulate expression of genes. Both of these goals are thought to be accomplished by altering the higher order of structure of chromosomes. In the nematode, C. elegans, two distinct condensin complexes perform these two functions. The mitotic condensin complex plays a role in chromosome segregation, while the dosage compensation complex regulates expression of genes on hermaphrodite X chromosomes to equalize X-linked gene dosage between the sexes. Our long-term goal is to decipher the molecular mechanism of worm dosage compensation and to determine how it relates to the mechanism of chromosome condensation. Biochemical purification of the dosage compensation complex led to the discovery of CPG-1, a new dosage compensation complex subunit, which also functions in chromosome segregation. To reveal mechanistic similarities between dosage compensation and chromosome segregation, we will use genetic, biochemical, and microscopic tools to analyze the ways in which CPG-1 can function during both processes. The second part of our proposed study will analyze the chromatin fiber of dosage compensated chromosomes. We found that HTZ-1, a histone variant of H2A, is depleted on dosage compensated X chromosomes as compared to autosomes. The goal of our proposed experiments is to decipher how this depletion contributes to dosage compensation. To do that we will study the relationship between the association of dosage compensation complex and HTZ-1 with the X chromosome, both at low resolution (immunofluorescence) and high resolution (chromatin immunoprecipitation). Healthy development of any organism depends on delivering the correct number of chromosomes to each cell and on each cell then turning on the right set of genes. Delivering the wrong number of chromosomes to cells, or inappropriately turning genes on or off, leads to developmental disorders or cancer. These functions of chromosomes depend largely on the structure and organization of DMA contained in them. A better knowledge of the basic biological principals of how chromosome structure determines gene activity levels and ensures proper chromosome segregation will be important to understanding both healthy development and the development of disease.