Matthews K R, Tschudi C, Ullu E
Department of Internal Medicine, Yale University School of Medicine, New Haven, Connecticut 06520-8022.
Genes Dev. 1994 Feb 15;8(4):491-501. doi: 10.1101/gad.8.4.491.
In trypanosomes, the generation of monocistronic mRNAs from polycistronic precursors is achieved via RNA processing, namely trans-splicing of the spliced leader sequence at the 5' end and cleavage/polyadenylation at the 3' end of the mRNA coding region. Recent evidence raised the intriguing possibility that these two reactions are coupled. To begin a dissection of the signals required for mRNA 5'-end and 3'-end formation and to uncover potential interactions between trans-splicing and polyadenylation, we mutagenized the intergenic region between the beta- and alpha-tubulin genes of Trypanosoma brucei. Block substitutions identified the pyrimidine-rich sequences at the alpha-tubulin 3'-splice-acceptor site as a major determinant for accurate trans-splicing downstream and 3'-end formation upstream. In addition to the utilization of cryptic 3'-splice sites, obliteration of the polypyrimidine tracts led to aberrant poly(A)+ site choice, even in the presence of the wild-type poly(A)+ site and neighboring sequences. Taken together, these results indicate that the polypyrimidine-rich sequences act as a bifunctional element that affects RNA processing both upstream and downstream from itself. This is consistent with the possibility that the polypyrimidine tract is recognized by both the trans-splicing and polyadenylation machineries, either sequentially or simultaneously.
在锥虫中，从多顺反子前体产生单顺反子mRNA是通过RNA加工实现的，即mRNA编码区5'端剪接前导序列的反式剪接以及3'端的切割/聚腺苷酸化。最近的证据提出了一种有趣的可能性，即这两个反应是偶联的。为了开始剖析mRNA 5'端和3'端形成所需的信号，并揭示反式剪接和聚腺苷酸化之间的潜在相互作用，我们对布氏锥虫β-微管蛋白基因和α-微管蛋白基因之间的基因间隔区进行了诱变。阻断取代确定了α-微管蛋白3'剪接受体位点富含嘧啶的序列是下游准确反式剪接和上游3'端形成的主要决定因素。除了利用隐蔽的3'剪接位点外，即使在存在野生型聚腺苷酸化位点和相邻序列的情况下，多嘧啶序列的缺失也会导致异常的聚腺苷酸化位点选择。综上所述，这些结果表明富含多嘧啶的序列作为一种双功能元件，影响其自身上下游的RNA加工。这与多嘧啶序列被反式剪接和聚腺苷酸化机制顺序或同时识别的可能性是一致的。
