Monocytes acquire prostate cancer specific chromatin conformations upon indirect co-culture with prostate cancer cells

Abstract

Background: Three-dimensional chromosome loop conformations are powerful regulators of gene expression. These chromosome conformations can be detected both in tumour and in circulating cells and have significant disease biomarker potential. We have recently detected specific chromosome conformations in circulating cells of patients with prostate cancer (PCa) which were similar to ones found in their primary tumours, however, the possibility of horizontal transfer of chromosome conformations was not studied previously.

Methods: Human monocytes (U937) were co-cultured in Boyden chambers through 0.4 uM membrane with or without PC-3 human PCa cells or their conditioned media and a custom DNA microarray for 900,000 chromosomal loops covering all coding loci and non-coding RNA genes was performed on each part of the co-culture system.

Results: We have detected 684 PC-3 cell-specific chromosome conformations across the whole genome that were absent in naïve monocytes but appeared in monocytes co-cultured with PC-3 cells or with PC-3-conditioned media. Comparing PC3-specific conformations to the ones we have previously detected in systemic circulation of high-risk PCa patients revealed 9 positive loops present in both settings.

Conclusions: Our results demonstrate for the first time a proof of concept for horizontal transfer of chromosome conformations without direct cell-cell contact. This carries high clinical relevance as we have previously observed chromatin conformations in circulating cells of patients with melanoma and PCa similar to ones in their primary tumours. These changes can be used as highly specific biomarkers for diagnosis and prognosis. Further studies are required to elucidate the specific mechanism of chromosome conformations transfer and its clinical significance in particular diseases.

Keywords: 3C; blood test; horizontal gene transfer; nucleome; prostate cancer.

Figure 1

Schematic representation of the transwell model and chromosomal conformation assay. (A) PC-3 cells were cultured at the bottom chamber for 48 hours, then U937 cells were placed in the top chamber and incubated for 24 hours. (B) Chromosome conformation assay: DNA is crosslinked, digested, ligated and new sequences (in places where loops were) are predicted using relevance machine vector. Loops presence is detected using DNA microarray (11).

Figure 2

Principal component analysis for the CCSs distribution between samples. Principal component analysis for the CCSs in PC-3 (A) and U937 (B) before and after co-cultures demonstrating a change in CCSs distribution.

Figure 3

Venn diagrams of CCSs overlap between treatment groups and high-risk PCa patients. (A–D). Venn diagrams indicating the number of overlapping CCSs between various groups.