Terns M P, Dahlberg J E, Lund E
Department of Biomolecular Chemistry, University of Wisconsin, Madison 53706.
Genes Dev. 1993 Oct;7(10):1898-908. doi: 10.1101/gad.7.10.1898.
We have identified cis-acting sequences that promote nuclear export of pre-U1 RNA injected into Xenopus oocyte nuclei. At least three elements, the 5' m7G cap, the 3'-terminal stem-loop structure, and sequences in the 5'-terminal 124 nucleotides, contribute to efficient export of this RNA. Both the 5' and 3' export signals can function separately and do so independently of the cap structure. Experiments using hybrid RNAs indicate that the 5' and 3' export sequences of U1 RNA are sufficient to direct export of the heterologous, otherwise nonexportable, U6 RNA. The absence of comparable export signals in U6 RNA appears to be responsible for its retention in the nucleus. Stability of the pre-snRNAs in the nucleus depends on the presence of both a 5' cap structure and a 3' base-paired stem. The 5' m7G cap is neither sufficient nor necessary for nuclear export. The m7G cap by itself did not promote export of U6 RNA or nonspecific small RNAs. Moreover, substitution of this cap with either an AppG cap or gamma-mppG cap did not eliminate export of either full-length or a "minimal" U1 RNA (lacking most of the internal U1 RNA sequences), but it reduced the rate of export by about two to threefold. However, in the absence of the 3' stem-loop, substitution of the m7G cap led to a greater decrease in export rate, underscoring the cooperative action of the three different export elements of pre-U1 RNA. The m7G cap analog, m7GpppG, selectively destabilized pre-U1 RNA within the nucleus. Thus, nuclear components that recognize the 5' m7G cap may be important for both the stability and the export of pre-U1 RNA.
我们已经鉴定出了顺式作用序列，这些序列可促进注射到非洲爪蟾卵母细胞核中的前体U1 RNA的核输出。至少有三个元件，即5' m7G帽、3'端茎环结构以及5'端124个核苷酸中的序列，有助于该RNA的有效输出。5'和3'输出信号均可单独发挥作用，且其作用独立于帽结构。使用杂交RNA进行的实验表明，U1 RNA的5'和3'输出序列足以指导异源的、否则无法输出的U6 RNA的输出。U6 RNA中缺乏类似的输出信号似乎是其滞留于细胞核中的原因。核内前体小核RNA的稳定性取决于5'帽结构和3'碱基配对茎的同时存在。5' m7G帽对于核输出既非充分条件也非必要条件。m7G帽本身并不能促进U6 RNA或非特异性小RNA的输出。此外，用AppG帽或γ-mppG帽替换该帽并不会消除全长或“最小”U1 RNA（缺乏大部分内部U1 RNA序列）的输出，但会使输出速率降低约两到三倍。然而，在没有3'茎环的情况下，m7G帽的替换导致输出速率下降幅度更大，这突出了前体U1 RNA的三种不同输出元件的协同作用。m7G帽类似物m7GpppG选择性地使核内的前体U1 RNA不稳定。因此，识别5' m7G帽的核成分可能对前体U1 RNA的稳定性和输出都很重要。
