Department of Zoology, University of Cambridge, Cambridge, United Kingdom.
Department of Genetics, University of Cambridge, Cambridge, United Kingdom.
PLoS Genet. 2022 Apr 28;18(4):e1010186. doi: 10.1371/journal.pgen.1010186. eCollection 2022 Apr.
At interphase, de-condensed chromosomes have a non-random three-dimensional architecture within the nucleus, however, little is known about the extent to which nuclear organisation might influence expression or vice versa. Here, using imprinting as a model, we use 3D RNA- and DNA-fluorescence-in-situ-hybridisation in normal and mutant mouse embryonic stem cell lines to assess the relationship between imprinting control, gene expression and allelic distance from the nuclear periphery. We compared the two parentally inherited imprinted domains at the Dlk1-Dio3 domain and find a small but reproducible trend for the maternally inherited domain to be further away from the periphery however we did not observe an enrichment of inactive alleles in the immediate vicinity of the nuclear envelope. Using Zfp57KO ES cells, which harbour a paternal to maternal epigenotype switch, we observe that expressed alleles are significantly further away from the nuclear periphery. However, within individual nuclei, alleles closer to the periphery are equally likely to be expressed as those further away. In other words, absolute position does not predict expression. Taken together, this suggests that whilst stochastic activation can cause subtle shifts in localisation for this locus, there is no dramatic relocation of alleles upon gene activation. Our results suggest that transcriptional activity, rather than the parent-of-origin, defines subnuclear localisation at an endogenous imprinted domain.
在间期，去凝聚的染色体在核内具有非随机的三维结构，然而，关于核组织在多大程度上可能影响表达或反之亦然，人们知之甚少。在这里，我们使用印迹作为模型，使用正常和突变的小鼠胚胎干细胞系中的 3D RNA 和 DNA 荧光原位杂交来评估印迹控制、基因表达和等位基因与核周缘的距离之间的关系。我们比较了 Dlk1-Dio3 区域的两个亲本遗传印迹域，发现母系遗传域离外周稍远，但我们没有观察到紧邻核膜的失活等位基因富集。使用 Zfp57KO ES 细胞，其携带从父系到母系的表观遗传转变，我们观察到表达的等位基因明显离外周更远。然而，在单个核内，靠近外周的等位基因与远离外周的等位基因同样有可能被表达。换句话说，绝对位置并不能预测表达。总之，这表明尽管随机激活可能导致该基因座的局部定位发生细微变化，但在基因激活时，等位基因不会发生明显的重新定位。我们的结果表明，转录活性而不是亲本来源决定了内源性印迹域的亚核定位。
