Royo Félix, Paz Nerea, Espinosa Luis, McQueen Philip G, Vellón Luciano, Parada Luis A
Cytogenomics, CIC bioGUNE-CIBEREHD, Par. Tec. Bizkaia Ed. 801 A, Derio, Spain.
Chromosoma. 2009 Dec;118(6):711-22. doi: 10.1007/s00412-009-0229-1. Epub 2009 Aug 1.
Eukaryotic genomes are highly organized within the cell nucleus. Genome organization not only implies the preferential positioning of genetic elements in the interphase nucleus but also the topographic distribution of biological processes. We have investigated the relationship between spatial organization and genome function in single cells. Myc, c-Met, Igf2r, Asb4, and Zac1 genes have the same radial distribution, but they are not positioned in close proximity with respect to each other. Three-dimensional mapping of their transcription sites uncovered a gene-specific pattern of relative positioning with respect to the nucleolus. We found that the Zac1 gene transcription preferentially occurs juxtaposed to the nucleolus, and that its mRNA accumulates at this site of transcription. Nucleoli isolation followed by qRT-PCR provided evidence for a physical interaction between Zac1 mRNA and the nucleolus. Actinomycin-D treatment induced disassembly of the nucleolus, loss of the RNA-FISH signal, and dramatic increase of the ZAC protein level. However, inhibition of RNA polymerase II had no effect over the Zac1 FISH signal and the protein expression. Induction of cell cycle arrest, which involves participation of the ZAC protein, provoked mRNA release from its retention site and protein synthesis. Our data demonstrate that Zac1 mRNA preferentially accumulates in close proximity to nucleoli within the cell nucleus. In addition, our results suggest a functional link between such spatial distribution and protein expression.
真核生物基因组在细胞核内高度有序组织。基因组组织不仅意味着遗传元件在间期核中的优先定位，还包括生物过程的拓扑分布。我们研究了单细胞中空间组织与基因组功能之间的关系。Myc、c-Met、Igf2r、Asb4和Zac1基因具有相同的径向分布，但它们彼此之间并未紧密定位。对其转录位点的三维图谱分析揭示了相对于核仁的基因特异性相对定位模式。我们发现Zac1基因转录优先发生在与核仁并列的位置，并且其mRNA在该转录位点积累。通过核仁分离后进行qRT-PCR，为Zac1 mRNA与核仁之间的物理相互作用提供了证据。放线菌素-D处理诱导核仁解体、RNA-FISH信号丧失以及ZAC蛋白水平显著增加。然而，抑制RNA聚合酶II对Zac1 FISH信号和蛋白表达没有影响。诱导涉及ZAC蛋白参与的细胞周期停滞，促使mRNA从其滞留位点释放并进行蛋白质合成。我们的数据表明，Zac1 mRNA优先在细胞核内核仁附近积累。此外，我们的结果表明这种空间分布与蛋白表达之间存在功能联系。
