Cotten M, Gick O, Vasserot A, Schaffner G, Birnstiel M L
Research Institute of Molecular Pathology, Vienna, Austria.
EMBO J. 1988 Mar;7(3):801-8. doi: 10.1002/j.1460-2075.1988.tb02878.x.
We have made a detailed molecular analysis of the reactions leading to the formation of mature 3' ends in mammalian histone mRNAs. Using two analytical protocols we have identified an essential sequence motif in the downstream spacer which is consistently present, albeit in diffuse form, mammalian histone genes. Tampering with this sequence element completely abolishes 3' processing. However, 3' cleavage in vitro, although at a very much reduced rate, can be detected when the conserved hairpin is deleted from histone precursor mRNAs. U7 snRNA, previously shown to be essential for the maturation of sea urchin histone messages, was isolated from murine cells and the sequence was determined. The approximately 63-nucleotide, trimethyl-G-capped, murine U7 snRNA possesses a sequence shown in the sea urchin U7 to be required for Sm-precipitability, and like the sea urchin U7, the 3' end of murine U7 is encased in a hairpin structure. The 5' sequence of murine U7 exhibits extensive sequence complementarity to the conserved downstream motif of the histone precursor. As expected, oligo-nucleotide-directed RNase H cleavage of this portion of murine U7 inhibits the in vitro processing reaction. These experiments identify a set of specific contacts between mammalian U7 and histone precursor RNA which is indispensable for the maturation reaction.
我们对哺乳动物组蛋白mRNA中成熟3'末端形成所涉及的反应进行了详细的分子分析。我们使用两种分析方法，在下游间隔区鉴定出一个必需的序列基序，该基序在哺乳动物组蛋白基因中始终存在，尽管呈弥散形式。对该序列元件进行破坏会完全消除3'加工。然而，当从组蛋白前体mRNA中删除保守的发夹结构时，虽然体外3'切割速率大幅降低，但仍可检测到。先前已证明U7 snRNA对海胆组蛋白mRNA的成熟至关重要，我们从鼠细胞中分离出U7 snRNA并确定了其序列。约63个核苷酸、带有三甲基-G帽的鼠U7 snRNA具有海胆U7中显示的Sm沉淀所需的序列，并且与海胆U7一样，鼠U7的3'末端被包裹在发夹结构中。鼠U7的5'序列与组蛋白前体的保守下游基序表现出广泛的序列互补性。正如预期的那样，针对鼠U7这部分的寡核苷酸导向的RNase H切割会抑制体外加工反应。这些实验确定了哺乳动物U7与组蛋白前体RNA之间的一组特定相互作用，这对于成熟反应是必不可少的。
