Amplified human MYC oncogenes localized to replicating submicroscopic circular DNA molecules.

Abstract

Amplification of genes can sometimes be detected by molecular hybridization but not by cytogenetic methods, suggesting that in some cases the units of amplification may be too small to be detected by light microscopy. The experiments reported here investigate whether submicroscopic amplification units are present in early passages of the human tumor cell lines HL-60 and COLO 320. The results show that such cells do contain submicroscopic, extrachromosomal, supercoiled circular molecules harboring MYC genes. The molecules in HL-60 are -250 kilobase pairs (kbp), while those in COLO 320 are 120-160 kbp. The extrachromosomal molecules in HL-60 are shown to replicate semiconservatively and approximately once in one cell cycle. We propose that these submicroscopic elements are precursors of double-minute chromosomes, the usual extrachromosomal manifestation of gene amplification, since both are structurally similar and replicate autonomously.The large number of oncogenes and drug-resistance genes subject to amplification indicates that many, possibly most, genomic regions are capable of undergoing regional increases in copy number (see refs. 1-3 for review). Cytogenetic analyses of many independently isolated cell lines containing different amplified genes show that amplified sequences localize to two types of abnormal chromosomal structures, referred to as double-minute chromosomes (DMs) and expanded chromosomal regions (also called homogeneously staining or abnormally banding regions). DMs are acentric extrachromosomal elements, which replicate approximately once per cell cycle and are believed to be circular (4, 5). Expanded chromosomal regions consist of amplification units arranged as both inverted and direct repeats (6-9). Size estimates for individual amplified units of different chromosomal regions range from 250 to >1000 kilobase pairs (kbp)

Amplification of cellular oncogenes: A predictor of clinical outcome in human cancer

“…We assume that the DNA released from the replication structure can be stabilized extrachromosomally and can replicate. This assumption would be consistent with the identification of populations of extrachromosomal replicating circular DNA molecules harboring amplified MYC genes (van Hoff et al, 1988), although the existence of such circular elements has not been formally established in neuroblastoma cells. It is quite possible that the extrachromosomal DNA may occupy structures that can be visualized as DMs.…”

Induction of Circles of Heterogeneous Sizes in Carcinogen-Treated Cells: Two-Dimensional Gel Analysis of Circular DNA Molecules

“…Extrachromosomal circles and gene amplification. Extrachromosomal circles were shown to be involved in early events of gene amplification both in cancerous mammalian cells (9,15,58,62,67,68,83,84,86,88,89) and in lower eukaryotes (6,23,26,29,31,34,37,61,63,82,87). These are usually large circles containing amplicons ranging in size from a few tens to a few hundreds of kilobase pairs, which are often organized in either an inverted or a direct order.…”

“…These molecules might not be detected under our experimental conditions. Alternatively, the small circles might evolve to larger molecules by secondary recombination events (9,15,28,58,67,68,83,84,86).…”

Cell-Free eccDNAs: A New Type of Nucleic Acid Component for Liquid Biopsy?

“…To unify the nomenclature, Moller et al has proposed to use extrachromosomal circular DNAs (eccDNAs) to describe all circular DNAs in eukaryotes (6). The eccDNAs have different sizes, including large circular DNAs such as episomes (10, 11) and double minute chromosomes (DMs) (12, 13), and the smaller circular molecules such as the small polydispersed DNA (spcDNAs) (14, 15) and microDNAs (5, 16). Episomes (from hundreds of kilobases to 1 megabase, submicroscopic) are considered as precursors of DMs (>1 megabase, microscopically visible).…”