PDF(Pubmed)
Abstract：
Chromatin exhibits non-random distribution within the nucleus being arranged into discrete domains that are spatially organized throughout the nuclear space. Both the spatial distribution and structural rearrangement of chromatin domains in the nucleus depend on epigenetic modifications of DNA and/or histones and structural elements such as the nuclear envelope. These components collectively contribute to the organization and rearrangement of chromatin domains, thereby influencing genome architecture and functional regulation. This study develops an innovative, user-friendly, ImageJ-based plugin, called IsoConcentraChromJ, aimed quantitatively delineating the spatial distribution of chromatin regions in concentric patterns. The IsoConcentraChromJ can be applied to quantitative chromatin analysis in both two- and three-dimensional spaces. After DNA and histone staining with fluorescent probes, high-resolution images of nuclei have been obtained using advanced fluorescence microscopy approaches, including confocal and stimulated emission depletion (STED) microscopy. IsoConcentraChromJ workflow comprises the following sequential steps: nucleus segmentation, thresholding, masking, normalization, and trisection with specified ratios for either 2D or 3D acquisitions. The effectiveness of the IsoConcentraChromJ has been validated and demonstrated using experimental datasets consisting in nuclei images of pre-adipocytes and mature adipocytes, encompassing both 2D and 3D imaging. The outcomes allow to characterize the nuclear architecture by calculating the ratios between specific concentric nuclear areas/volumes of acetylated chromatin with respect to total acetylated chromatin and/or total DNA. The novel IsoConcentrapChromJ plugin could represent a valuable resource for researchers investigating the rearrangement of chromatin architecture driven by epigenetic mechanisms using nuclear images obtained by different fluorescence microscopy methods.
摘要：
染色质在细胞核内表现出非随机分布，被排列成在整个核空间中空间组织的离散域。核中染色质结构域的空间分布和结构重排都取决于DNA和/或组蛋白以及诸如核膜的结构元件的表观遗传修饰。这些成分共同有助于染色质结构域的组织和重排，从而影响基因组结构和功能调控。这项研究开发了一种创新，用户友好，基于ImageJ的插件，叫做IsoConcentraChromJ，旨在定量描绘同心模式中染色质区域的空间分布。IsoConcentraChromJ可应用于二维和三维空间中的定量染色质分析。用荧光探针进行DNA和组蛋白染色后，使用先进的荧光显微镜方法获得了核的高分辨率图像，包括共聚焦和受激发射损耗（STED）显微镜。IsoConcentraChromJ工作流程包括以下顺序步骤：核分割，阈值，掩蔽,归一化，和三分法，具有指定的二维或三维采集比率。IsoConcentraChromJ的有效性已通过实验数据集得到验证和证明，这些数据集包括前脂肪细胞和成熟脂肪细胞的细胞核图像，包括2D和3D成像。结果允许通过计算乙酰化染色质相对于总乙酰化染色质和/或总DNA的特定同心核面积/体积之间的比率来表征核结构。新的IsoConcentrapChromJ插件可以代表研究人员使用通过不同荧光显微镜方法获得的核图像研究表观遗传机制驱动的染色质结构重排的宝贵资源。
