The functions of DNA topoisomerase and genes regulated by DNA topology in differentiation.

在细胞分化中，各种基因的表达是通过改变DNA拓扑结构来调节的，而DNA拓扑结构受到DNA拓扑异构酶I和II的紧密调控。我们发现了两个在与拓扑异构酶抑制剂孵育早期就被抑制的基因。其中一个与hnRNPA1高度同源，hnRNPA1调节选定转录本的剪接或稳定mRNAs。另一个是一个新基因，命名为TIS，全长约21kb，包括11个外显子，以单拷贝形式存在。它在大多数小鼠组织中都有表达，但其功能尚不清楚。然后，我们分析了拓扑异构酶抑制剂(依托泊苷)、反式维甲酸(Tra)和二丁酰环腺苷(Bt2cAMP)等分化药物对人涎腺肿瘤细胞(HSG)中拓扑异构酶和TIS基因表达的影响。拓扑异构酶I的表达被trA抑制，但不被bt2cAMP抑制。拓扑异构酶I的表达被tra抑制，但被bt2cAMP抑制的程度较小。拓扑异构酶IIβ…的水平更多的是低的，没有改变。这些结果表明，拓扑异构酶IIα的下调可能通过改变DNA的拓扑结构来阻止细胞的增殖和启动分化。药物处理后早期TIS基因表达降低，提示TIS基因参与了细胞分化。近年来，已知TIS在肿瘤转化、细胞因子诱导的T细胞活化、肿瘤增殖和细胞凋亡中发挥一定的重要作用，诱导细胞凋亡是由于涎腺腺泡细胞等多种细胞的终末分化。拓扑异构酶II抑制剂依托泊苷诱导涎腺腺泡细胞和HSG细胞发生凋亡。已知在凋亡细胞中，PKCδ被caspase-3所切割。我们在小鼠唾液腺中发现了一种对半胱氨酸天冬氨酸氨基转移酶-3不敏感的新的PKCδ亚型。这些发现表明，一个新的蛋白激酶Cδ亚型可能在涎腺细胞的终末分化过程中起着重要的作用，需要进一步研究拓扑异构酶和TIS基因在同质分化细胞中的作用。较少

In cellular differentiation, various gene expression is regulated by altering DNA topology which is tightly modulated by DNA topoisomerase I and II.We identified two genes that were suppressed early in the incubation with topoisomerase inhibitors. One was highly homologous to hnRNPA1 which modulates splicing of selected transcripts or stabilizes mRNAs. The other was a novel gene, named as TIS, which spanned about 21 kb including 11 exons and was present as a single copy. It was expressed in most mouse tissues, but its function was unknown. Then, we analyzed the roles of topoisomerases and TIS gene in human salivary gland tumor cells (HSG) treated by differentiating agents such as topoisomerase inhibitor (etoposide), trans-retinoic acid (tRA) or dibutyryl cyclic AMP (bt2cAMP). Expression of topoisomerase I was reduced at the early state by tRA treatment but not by bt2cAMP.Expression of topoisomerase I was decreased by tRA, but at a lesser extent by bt2cAMP.The level of topoisomerase IIβ … More was low and was not altered. The results suggest that downregulation of topoisomerase IIα involves in arrest of cellular proliferation and initiation of differentiation through alterations in DNA topology. TIS gene expression was reduced at the early stage after the treatment of the agents, suggesting the involvement of TIS gene in differentiation. Recently, TIS has been known to play certain important role in neoplastic transformation, cytokine-induced T cell activation, neoplastic proliferation, and apoptosis.Apoptosis is induced as terminal differentiation in various cells inculuding salivary gland acinar cells. Salivary gland acinar cells and HSG cells underwent apoptosis by a topoisomerase II inhibitor, etoposide. PKCδ is known to be cleaved by caspase-3 in apoptotic cells. We found a novel isoform of PKCδ which was insensitive to caspase-3 in mouse salivary glands. These findings suggest that a novel PKCδ isoform may have some important role in terminal differentiation of salivary gland cells.Further studies are required to elucidate the role of topoisomerases and TIS gene using homogeneously differentiating cells as a model. Less