Department of Physics and Astronomy "Ettore Majorana", University of Catania, Catania, Italy.
Nanoscopy and NIC@IIT, CHT Erzelli, Istituto Italiano di Tecnologia, Genoa, Italy.
Commun Biol. 2024 Oct 4;7(1):1260. doi: 10.1038/s42003-024-06972-2.
To understand how oncogenes affect genome organization, it is essential to visualize fundamental processes such as DNA replication and transcription at high resolution in intact cells. At the same time, it is important to determine the progression of the cell along the cell cycle, as cell cycle regulation is crucial for the control of cell proliferation and oncogenesis. Here, we present a super-resolution imaging-based method to analyze single cell nuclei sorted according to specific phases of the cell cycle. The sorting is based on the evaluation of the number and the intensity of pixels in the replication foci image and the colocalization analysis is based on image cross-correlation spectroscopy (ICCS). We evaluate the colocalization between replication and transcription, at different cell cycle phases, in a model of PML-RARα oncogene activation. We find that colocalization between replication and transcription is higher in cells in early S phase compared to cells in middle and late S phase. When we turn on the PML-RARα oncogene, this colocalization pattern is preserved but we detect an increase of colocalization between replication and transcription in the early S phase which points to an effect of the PML-RARα oncogene on the coordination between replication and transcription.
为了了解癌基因如何影响基因组的组织，必须在完整细胞中高分辨率地可视化基本过程，如 DNA 复制和转录。同时，确定细胞在细胞周期中的进展也很重要，因为细胞周期调控对于控制细胞增殖和癌变至关重要。在这里，我们提出了一种基于超分辨率成像的方法，用于分析根据细胞周期特定阶段分选的单个细胞核。这种分选是基于对复制焦点图像中的像素数量和强度的评估，而共定位分析则基于图像互相关光谱学 (ICCS)。我们评估了在 PML-RARα 癌基因激活模型中，不同细胞周期阶段的复制和转录之间的共定位。我们发现，与中晚期 S 期相比，早期 S 期细胞中复制和转录之间的共定位更高。当我们开启 PML-RARα 癌基因时，这种共定位模式得以保留，但我们在早期 S 期检测到复制和转录之间的共定位增加，这表明 PML-RARα 癌基因对复制和转录之间的协调有影响。
