Cancer cells run rings around their normal counterparts in terms of proliferation, survival, migration, and aggressivity to devastating effect, but how do cancer cells get themselves “in shape” to allow for these strenuous activities? Previously, researchers led by Howard Y. Chang (Stanford University), Bing Ren, Vineet Bafna, and Paul S. Mischel (University of California at San Diego, USA) established that cancer cells possess huge numbers of circular oncogene-containing extrachromosomal DNA (ecDNA) particles and, in their recent study, the team evaluated how circular ecDNA particles shape the transcriptional profiles of cancer cells and explored the possible involvement of altered chromatin organization.
So, we gave the lab a “ring” to say “halo” and to discuss their “revolution”-ary new study; here are the details of how their “hoop”ful examination into the chromatin organization of circular ecDNA shaped up:
- A combination of DNA sequencing, optical
three-dimensional structured illumination microscopy, and both
scanning and transmission electron microscopy-based analyses confirmed the
circular nature of ecDNAs from cancer cells
- Amplicon sizes range from 168 kb to 5 Mb
- Distinguishing single nucleotide polymorphisms
highlight the increased expression of ecDNA-coded oncogenes derived from cancer
cells and clinical tumor samples when compared to their linear
- ecDNA-encoded genes, which include EGFR, MYC, CDK4, and MDM2, are among the most highly expressed genes in the tumor transcriptome
- While higher copy number of ecDNA drives increased oncogene transcription, the elevated transcription of ecDNA-coded oncogene transcripts even when normalized to copy number suggests that chromatin organization may also play a role
- A vast range of techniques used to
quantitatively assess chromatin state provided evidence that ecDNA possesses very
highly accessible chromatin, possibly owing to a lack of chromatin compaction
- Histone modification analysis by immunofluorescence and chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) revealed the presence of active, and the lack of repressive, histone modifications in cancer cell-derived ecDNA
- Chromatin accessibility and nucleosome mapping analyses using assay for transposase-accessible chromatin using sequencing (ATAC-seq) and micrococcal nuclease digestion and sequencing (MNase-seq) confirmed the packaging of ecDNA into nucleosome units; however, ecDNA packaging lacks the tight higher-order compaction of chromosomal DNA, leading to higher chromatin accessibility
- Additionally, ATAC with visualization (ATAC-see) directly confirms the increase in the accessibility of ecDNA chromatin
- Finally, the authors employ proximity ligation-assisted ChIP-seq (PLAC-seq) and circular chromosome conformation capture combined with high-throughput sequencing (4C-seq) to establish that a notably increased number of ultra-long-range interactions occurs on ecDNA suggesting that it can affect distal gene expression
“By showing that ecDNA is circular, then elucidating its epigenetic organization, we demonstrate something very powerful,” said Mischel, co-senior author of the study. “This unique shape in human cancer cells is quite unlike normal human DNA. It really shines a new light onto the 3D organization of the screwed-up cancer genome and epigenome, which now provides a mechanistic basis for understanding why certain tumor cells are so aggressive.”
Overall, the authors provide the potential mechanism by which cancer cells get “in shape” by showing that circular ecDNA possesses a highly accessible chromatin landscape that enhances the expression of resident oncogenes – can we use this knowledge to shape the development of novel and effective anti-cancer therapeutics?
For now, “donut” go anywhere! Circle around, settle in for the night, and read more at Nature, November 2019.