Knop Katarzyna, Stepien Agata, Barciszewska-Pacak Maria, Taube Michal, Bielewicz Dawid, Michalak Michal, Borst Jan W, Jarmolowski Artur, Szweykowska-Kulinska Zofia
Department of Gene Expression, Institute of Molecular Biology and Biotechnology, Faculty of Biology, Adam Mickiewicz University, Umultowska 89, Poznan 61-614, Poland.
Department of Molecular and Cellular Biology, Institute of Molecular Biology and Biotechnology, Faculty of Biology, Adam Mickiewicz University, Umultowska 89, Poznan, 61-614, Poland.
Nucleic Acids Res. 2017 Mar 17;45(5):2757-2775. doi: 10.1093/nar/gkw895.
Arabidopsis, miR402 that is encoded within the first intron of a protein-coding gene At1g77230, is induced by heat stress. Its upregulation correlates with splicing inhibition and intronic proximal polyA site selection. It suggests that miR402 is not processed from an intron, but rather from a shorter transcript after selection of the proximal polyA site within this intron. Recently, introns and active 5' splice sites (5'ss') have been shown to stimulate the accumulation of miRNAs encoded within the first exons of intron-containing MIR genes. In contrast, we have observed the opposite effect of splicing inhibition on intronic miR402 production. Transient expression experiments performed in tobacco leaves revealed a significant accumulation of the intronic mature miR402 when the 5'ss of the miR402-hosting intron was inactivated. In contrast, when the miR402 stem-loop structure was moved into the first exon, mutation of the first-intron 5'ss resulted in a decrease in the miRNA level. Thus, the 5'ss controls the efficiency of miRNA biogenesis. We also show that the SERRATE protein (a key component of the plant microprocessor) colocalizes and interacts with several U1 snRNP auxiliary proteins. We postulate that SERRATE-spliceosome connections have a direct effect on miRNA maturation.
拟南芥中，位于蛋白质编码基因At1g77230第一个内含子内编码的miR402受热胁迫诱导。其上调与剪接抑制和内含子近端聚腺苷酸化位点选择相关。这表明miR402不是从内含子加工而来，而是在该内含子内选择近端聚腺苷酸化位点后从较短的转录本加工而来。最近，内含子和活跃的5'剪接位点（5'ss'）已被证明可刺激含内含子的MIR基因第一个外显子内编码的miRNA的积累。相比之下，我们观察到剪接抑制对内含子miR402产生有相反的影响。在烟草叶片中进行的瞬时表达实验表明，当miR402所在内含子的5'ss失活时，内含子成熟miR402会大量积累。相反，当miR402茎环结构移至第一个外显子时，第一个内含子5'ss的突变导致miRNA水平降低。因此，5'ss控制着miRNA生物合成的效率。我们还表明，锯齿蛋白（植物微处理器的关键组成部分）与几种U1 snRNP辅助蛋白共定位并相互作用。我们推测锯齿蛋白-剪接体连接对miRNA成熟有直接影响。
