DNA replication timing and other coordinately regulated properties of chromatin organization are disrupted at oncogenic loci in cancer. Pleiotrophin (PTN) is an oncogene expressed in all malignant gliomas and is transcriptionally regulated during neural differentiation coincident with changes to its replication timing. Such links between chromatin organization and oncogene function underscore the need for mechanistic understanding of how chromatin organization is regulated. The long-term goal is to understand how cells organize chromatin to regulate genomic function to identify and correct the loss of proper organization in cancer. The immediate objective of this application is to define the role and nature of cis elements in the regulation of chromatin organization at the PTN locus. The central hypothesis is that the chromosomal domain in which PTN resides contains DNA elements that allow the domain to be recognized and regulated as an individual functional unit that corresponds to the region that changes replication timing. Preliminary data produced by the applicant and others lead to the formulation of this hypothesis. The rationale that underlies the proposed research is that understanding the role and nature of cis regulation of the PTN chromosomal domain will allow for hypothesis-driven approaches to identify the trans factors responsible for organizing chromatin in the nucleus and will provide insight as to how oncogenes are activated during oncogenesis. The central hypothesis will be tested by pursuing two specific aims: 1) Isolate minimal units of replication timing regulation at the PTN locus;and 2) Determine the role of replication timing in the functional regulation of PTN. To achieve these aims, single-copy segments of the PTN domain will be integrated into a different chromosomal domain with opposite replication timing. Segments that maintain replication-timing regulation will be reduced to minimal units of regulation for the first aim. Under the second aim, PTN promoter activity in segments that do not maintain replication-timing regulation will be compared to activit at the endogenous locus. The expected contributions of the proposed research are to reveal the nature of cis elements regulating chromatin organization and determine their role in the regulation of gene function. These contributions will be significant because identifying regulatory elements is our first step toward uncovering the mechanisms governing cell-type specific chromatin organization and its links to cancer.
The proposed research is relevant to public health because advances in our understanding of the mechanism whereby cells differentially regulate DNA function provide insight as to how that regulation is undermined in cancer. This project is relevant to the mission of National Cancer Institute because understanding the regulation of oncogene function is critical to understanding how oncogenes are activated to cause cancer and what measures should be taken to prevent or reverse their activation in patients.
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