Functional components of a mammalian origin of DNA replication and the possible role of an origin in human Fragile X Mental Retardation
Gray, Steven James
This project was concerned with the functional components of mammalian DNA replication origins and how the misuse of a start site for DNA replication at the FMR1 locus might contribute to human Fragile X Syndrome. In the first part of this dissertation, I identified a novel origin of DNA replication near the CGG repeats at the human Fragile X Mental Retardation (FMR1) gene promoter. Expansion of these repeats leads to the epigenetic chromosome modifications that cause Fragile X Syndrome. The experiments described in this dissertation suggest that the position of the FMR1 origin favors contraction of the CGG repeats, thus providing a mechanism to avoid repeat expansion. This model predicts that a change in origin usage accompanies repeat expansion, and I discussed how this could occur. In the second part of this dissertation, I examined the requirement of DNA sequence elements in a mammalian origin to direct DNA replication to start at specific chromosomal sites. In particular, I studied the role of a dinucleotide repeat (DNR) sequence element in the activity of the Chinese hamster dihydrofolate reductase origin beta. The DNR element could be functionally replaced with two different transcriptional elements. This result suggests that DNR shares a functional role with these elements, and we speculate that this role may be to create the proper chromatin environment for recruitment and action of other replication factors to initiate replication.