double minute chromosomes in ovarian cancer
Sequence Level Analysis of Double Minute Chromosomes in the Ovarian Cancer Cells Yields Insights into Their Complicated Molecular Architecture
Double minute chromosomes (DMs), the autonomously replicating extra-chromosomes, are frequently associated with gene amplification in tumors. However, their molecular architecture and the underlying mechanisms leading to their formation and function are largely unknown.In our study, the rearrangements illustrated by genetic sequencing suggest that DM is formed by fusion of amplified or non-amplified DNA fragments. The complicated rearrangement patterns presented in current study suggest that the generation and evolution of DMs is attributable to the action of the non-homologous end joining and single strand annealing pathways on the assembly of these chromosomal fragments. Furthermore, we found for the first time that retrotransposed pseudogene EIF5A2 was associated with DMs in the ovarian cancer cell line UACC-1598. This study on sequencing purified DMs provides a new perspective on the architecture of DMs, highlighting the complex molecular structure and the possible events for their formation and evolution of DMs.
We established optimal conditions for two-dimensional pulsed field gel electrophoresis and purified DMs from ovarian cancer cell line UACC-1598. The DMs in the cancer cells are heterogeneous in size (1–3 Mb), and the purified DNA molecules were confirmed as intracellular DMs using fluorescence in situ hybridization on metaphase chromosome. The entire genetic landscape of DMs was determined by second-generation sequencing technology. We found that DMs arised from multiple chromosomal regions (including four major amplified fragments from 1q21.2, 2p24.3, 3q26.2 and 6p22.3), and were heavily rearranged with 157 rearrangement junctions.