Activation of a mammalian origin of replication by chromosomal rearrangement.
Activation of a mammalian origin of replication by chromosomal rearrangement.
复制标题
通过染色体重排激活哺乳动物复制起点。
DOI：
10.1128/mcb.12.6.2804-2812.1992
复制
发表时间：
1992
影响因子：
5.3
通讯作者：
Hamlin,JL
中科院分区：
文献类型：
--
作者：
Leu,TH;Hamlin,JL
抗甲氨蝶呤的中国仓鼠细胞系DC3F/A3-4K(A3/4K)至少含有两种突出的二氢叶酸还原酶扩增子类型。I型扩增片段的长度至少为650kb，占总扩增片段的80%，但其末端结构尚未确定。第二类序列约占扩增产物的20%，全长450kb，以头对头和尾对尾交替排列。在先前对扩增片段较小的CHEC400品系的研究中，已经证明在二氢叶酸还原酶基因的下游存在一个复制起始点(οri-β/οri-γ)。在最近对A3/4K细胞较大扩增片段的研究中，我们发现了一个额外的起始点(ORI-α)，位于οri-β/οri-γ上游约240kb。有趣的是，体内标记实验表明，复制叉只在下游方向与Ori-α背道而驰。这一发现表明，Ori-Cx是一个单向起源，或者终点位于Ori-α的上游。然而，在本研究中，我们发现Ori-α实际上非常接近A3/4K细胞中微小的II型扩增片段之间的头到头的回文连接序列；此外，Ori-α在S早期的II型扩增片段中是活跃的，但在缺乏这种回文连接的较大的I型序列中不是。这是在哺乳动物细胞中首次直接证明了染色体重排可以激活隐蔽的起源，推测是通过删除负调控元件或通过创造更有利的染色体启动环境来实现的。
The methotrexate-resistant Chinese hamster cell line DC3F/A3-4K (A3/4K) contains at least two prominent dihydrofolate reductase amplicon types. The type I amplicons, constituting ~80% of the total, are at least 650 kb in length, but the endpoints have not yet been characterized. The type II sequences represent ~20% of amplicons, are 450 kb in length, and are arranged as alternating head-to-head and tail-to-tail repeats. In previous studies on the CHOC 400 line, in which the amplicons are much smaller, a replication initiation locus (οri-β/οri-γ) has been shown to reside downstream from the dihydrofolate reductase gene. In a more recent study on the larger amplicons of A3/4K cells, we detected an additional initiation locus (ori-α) lying ~240 kb upstream from οri-β/οri-γ. Interestingly, in vivo labelling experiments suggested that replication forks diverge from ori-α only in the downstream direction. This finding suggested either that ori-cx is a unidirectional origin or that a terminus lies immediately upstream from ori-α. However, in this study, we show that ori-α is actually very close to the head-to-head palindromic junction sequence between the minor type II amplicons in A3/4K cells; furthermore, ori-α is active in the early S period in the type II amplicons but not in the larger type I sequences that lack this palindromic junction. This is the first direct demonstration in mammalian cells that a cryptic origin can be activated by chromosomal rearrangement, presumably by deleting negative regulatory elements or by creating a more favorable chromosomal milieu for initiation.