The instructions that guide the normal operations of a cell are contained within its DNA. In each cell the DNA is packaged into structures called chromosomes. Cells constantly divide to produce new cells that replace cells that are worn-out. During the process of division, it is of utmost importance that the chromosomes remain intact and that the correct number of chromosomes is distributed to each new cell. Changes in chromosome structure and number within cells can disrupt basic cellular functions leading to cancer. It is also essential that the reproductive cells, eggs and sperm, have the correct number of chromosomes and that these chromosomes also have the correct structure. In the event that the chromosome structure or number is compromised (aneuploidy) in eggs or sperm, the resulting offspring may fail to develop properly. Cancer treatments often have negative side effects including secondary tumors and infertility. The DNA disentangling enzyme Topoisomerase II (Topo II) is an important target of several anti-cancer drugs. The normal function of this enzyme is to unknot and untangle DNA as cells go through mitosis. Topoisomerase II is present in many different cells including cells that are undergoing the specialized cell division of meiosis. The roles of Topo II have been extensively studied in mitosis, but much less is known about its normal function in the meiotic cell cycle. Our long-term goal is to understand the molecules and systems that ensure that each cell receives the correct number of chromosomes during meiosis. In this proposal, we will investigate how Topo II interacts with the known players involved in maintaining chromosome structure during meiosis (Aim 1) and we will identify the components that make sure that Topoisomerase II operates at the correct time and place during this cell division (Aim 2). Additionally, both of these aims will address how Topo II differentially contributes to meiosis in male and female germ lines. Utilizing a temperature-sensitive allele of topo II [C. elegans top-2(it7ts)] as a unique tool the proposed research will address the following goals:
In Specific Aim 1, a combination of live cell imaging and cytological approaches will be used to characterize the role of top-2 in male and female meiosis and the relationship between TOP-2, the meiotic cohesins, and Aurora B kinase.
Specific Aim 2 will elucidate the mechanisms of TOP-2 regulation during male and female meiosis using a proteomics approach. Ultimately, this research will contribute to our understanding of how chromosomes are properly segregated to prevent the formation of aneuploid gametes that lead to birth defects. In the future, we hope this research may lead to the identification of potential therapeutic targets to prevent the formation of aneuploid gametes.
The formation of gametes with the ?wrong? number of chromosomes (aneuploidy) is a key contributing factor to infertility and birth defects in humans (30% of miscarriages, ~4% of stillborns, and ~0.3% of liveborns). The DNA disentangling enzyme Topoisomerase II is an enzyme that unknots and untangles DNA during normal cellular processes, but how this important enzyme is regulated during the cell cycle that a makes sperm and eggs (meiosis) is unknown. The proposed research will provide fundamental insights into the functions and regulation of Topoisomerase II in meiotic chromosome structure and segregation, which may lead to the identification of potential therapeutic targets to prevent the formation of aneuploid gametes.
