The Role of a Scaffold Attachment Factor in Nuclear Assembly and Architecture

正在进行的项目集中在非洲爪哇卵提取物中特定的DNA结合蛋白SAF-A(支架附着因子A)在细胞核组装中的作用。在这些提取物中，细胞核在添加的DNA或染色质周围自发形成。在赠款的第一阶段，我们使用显微镜和生化方法研究了SAF-A在核组装过程中的命运。我们可以证明，这种蛋白质与体外组装的细胞核中的不溶性核支架有关。随后的实验将研究特定的DNA结合对蛋白质定位的作用，并将表明蛋白质是否是组装或维护核支架所必需的。在此背景下，我们还将绘制负责SAF-A与核支架其他蛋白质成分相互作用的蛋白质结构域以及SAF-A多聚体的自组装。另一个项目分析了SAF-A对体外组装在人类有丝分裂染色体周围的染色体区域结构的作用。我们希望这些实验将提供一个详细的洞察力，以了解SAR结合蛋白对细胞核结构的作用。

The ongoing project is focussed on the role of the SAR-Specific DNA binding protein SAF-A (Scaffold Attachment Factor A) in the assembly of nuclei in extracts from Xenopus laevis eggs. In these extracts, nuclei form spontaneously around added DNA or chromatin. In the first period of the grant, we used microscopical and biochemical methods to investigate the fate of SAF-A during nuclear assembly. We could show that the protein associates with the insoluble nuclear scaffold in in vitro-assembled nuclei. Subsequent experiments will now investigate the role of the specific DNA binding for the localization ot the protein, and will show if the protein is necessary for the assembly or maintenance of a nuclear scaffold. In this context, we will also map the protein domain responsible for interaction of SAF-A with other protein components of the nuclear scaffold and for the self-assembly of SAF-A multimers. An additional project analyzes the role of SAF-A for the structure of chromosome territories assembled in vitro around human mitotic chromosomes. We expect that these experiments will provide a detailed insight into the role of a SAR binding protein for the architecture of the nucleus.