Possible Specific Role Of Triplex And Hairpin DNA Structures In Causing Microsatellite Instability Through Different Modes Of Replication Cycle
Open Access
- Author:
- Law, Qiao Kai
- Area of Honors:
- Biotechnology
- Degree:
- Bachelor of Science
- Document Type:
- Thesis
- Thesis Supervisors:
- Maria Krasilnikova, Thesis Supervisor
David Scott Gilmour, Thesis Honors Advisor
Dr. Wendy Hanna-Rose, Faculty Reader - Keywords:
- Possible Specific Role of triplex hairpin DNA secondary structur
- Abstract:
- Microsatellite is a tandem array of repeated nucleotides that consists of six or more repeated motif units. They are present all over the genome and are relatively unstable compared to normal DNA sequences. When microsatellite exceeds the threshold length, it can undergo large-scale expansions/contractions, which is often referred to as microsatellite instability. Many microsatellite repeats, especially trinucleotides, have been linked to over 20 types of neurological disorders and cancers. However, the mechanism of microsatellites instability in the genome that sometimes leads to the diseases is still enigmatic. Here we studied the effect of di- and tetra-nucleotides repeats in the first replication cycle as well as in the subsequent replication cycles. Replication of SV40-origin based plasmid transiently transfected in 293A or Cos-1 cells, served as model system for the first and subsequent replication cycles respectively. Plasmid replication was analyzed using two-dimensional gel electrophoresis to visualize the replication effects of different di- and tetranucleotide repeats in two different replication modes. We showed that the first replication cycle was more prone to stalling by triplex-forming repeats, while the subsequent replication cycle was mostly stalled by the repeats with hairpin-forming capacity. The replication stalling capacity of different repeats that we analyzed was consistent with the microsatellite mutability rates observed in the human-chimpanzee genomic evolution, indicating that replication stalling may be an important factor driving the instability of those repeats. It is interesting that triplex-forming, and not hairpin-forming potential of repeats, had a better correlation with the previously observed mutability rate in tetranucleotides. However, for dinucleotides, it seemed that hairpin formation correlated better with instability while triplexes had only a limited role. We suggest that triplex structure may have an impact on genomic instability when chromatin structure is loose, which occurs in the first replication cycle of transfected DNA. Comparison between the replication stalling effects of microsatellite in different replication mode with the mutability in chimpanzee-human evolution may indicate the specific role of different DNA secondary structures in causing microsatellite instability.