Wilson G M, Sun Y, Sellers J, Lu H, Penkar N, Dillard G, Brewer G
Department of Microbiology and Immunology, Wake Forest University School of Medicine, Winston-Salem, North Carolina 27157, USA.
Mol Cell Biol. 1999 Jun;19(6):4056-64. doi: 10.1128/MCB.19.6.4056.
The A+U-rich RNA-binding factor AUF1 exhibits characteristics of a trans-acting factor contributing to the rapid turnover of many cellular mRNAs. Structural mapping of the AUF1 gene and its transcribed mRNA has revealed alternative splicing events within the 3' untranslated region (3'-UTR). In K562 erythroleukemia cells, we have identified four alternatively spliced AUF1 3'-UTR variants, including a population of AUF1 mRNA containing a highly conserved 107-nucleotide (nt) 3'-UTR exon (exon 9) and the adjacent downstream intron (intron 9). Functional analyses using luciferase-AUF1 3'-UTR chimeric transcripts demonstrated that the presence of either a spliceable or an unspliceable intron 9 in the 3'-UTR repressed luciferase expression in cis, indicating that intron 9 sequences may down-regulate gene expression by two distinct mechanisms. In the case of the unspliceable intron, repression of luciferase expression likely involved two AUF1-binding sequences, since luciferase expression was increased by deletion of these sites. However, inclusion of the spliceable intron in the luciferase 3'-UTR down-regulated expression independent of the AUF1-binding sequences. This is likely due to nonsense-mediated mRNA decay (NMD) owing to the generation of exon-exon junctions more than 50 nt downstream of the luciferase termination codon. AUF1 mRNA splice variants generated by selective excision of intron 9 are thus also likely to be subject to NMD since intron 9 is always positioned >137 nt downstream of the stop codon. The distribution of alternatively spliced AUF1 transcripts in K562 cells is consistent with this model of regulated AUF1 expression.
富含A+U的RNA结合因子AUF1具有反式作用因子的特征，有助于许多细胞mRNA的快速周转。AUF1基因及其转录的mRNA的结构图谱显示在3'非翻译区（3'-UTR）内存在可变剪接事件。在K562红白血病细胞中，我们鉴定出四种可变剪接的AUF1 3'-UTR变体，包括一群含有高度保守的107个核苷酸（nt）的3'-UTR外显子（外显子9）和相邻下游内含子（内含子9）的AUF1 mRNA。使用荧光素酶-AUF1 3'-UTR嵌合转录本进行的功能分析表明，3'-UTR中可剪接或不可剪接的内含子9的存在顺式抑制荧光素酶表达，表明内含子9序列可能通过两种不同机制下调基因表达。对于不可剪接的内含子，荧光素酶表达的抑制可能涉及两个AUF1结合序列，因为删除这些位点会增加荧光素酶表达。然而，荧光素酶3'-UTR中包含可剪接内含子会下调表达，而与AUF1结合序列无关。这可能是由于在荧光素酶终止密码子下游超过50 nt处产生外显子-外显子连接导致的无义介导的mRNA衰变（NMD）。因此，通过选择性切除内含子9产生的AUF1 mRNA剪接变体也可能受到NMD的影响，因为内含子9总是位于终止密码子下游>137 nt处。K562细胞中可变剪接的AUF1转录本的分布与这种调节AUF1表达的模型一致。
