Systematic profiling of poly(A)+ transcripts modulated by core 3' end processing and splicing factors reveals regulatory rules of alternative cleavage and polyadenylation.
Systematic profiling of poly(A)+ transcripts modulated by core 3' end processing and splicing factors reveals regulatory rules of alternative cleavage and polyadenylation.
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DOI：
10.1371/journal.pgen.1005166
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发表时间：
2015-04
期刊：
影响因子：
4.5
通讯作者：
Tian B
中科院分区：
文献类型：
--
作者：
Li W;You B;Hoque M;Zheng D;Luo W;Ji Z;Park JY;Gunderson SI;Kalsotra A;Manley JL;Tian B
选择性裂解和聚腺苷化（APA）导致mRNA亚型含有不同的3 ‘非翻译区（3 ’ utr）和/或编码序列。核心裂解/聚腺苷酸化（C/P）因子如何调节APA尚不清楚。通过siRNA敲低和深度测序，我们发现几个C/P因子可以在3'UTR-APA中发挥重要作用。Pcf11和Fip1促进近端聚(A)位点的使用，而CFI-25/68、PABPN1和PABPC1促进远端聚(A)位点的使用。CFI-25/68或Fip1调控的pAs存在明显的顺式元件偏倚，pAs之间的距离在APA调控中起重要作用。此外，内含子pAs受到剪接因子的大量调控，其中U1主要抑制基因5 '端附近内含子中的C/P事件，U2主要抑制具有高效剪接特征的内含子中的C/P事件。此外，PABPN1抑制转录起始位点（TSS）附近pAs转录本的表达，这一特性可能与其在RNA降解中的作用有关。最后，我们发现由C/P因子调控的APA事件组在细胞分化和发育过程中也受到不同趋势的调节。总之，我们的研究结果支持一个APA编码，其中在给定的细胞环境中，APA事件受到许多参数的调节，包括相对于TSS的位置、剪接环境、竞争pAs之间的距离、周围顺式元件和核心C/P因子的浓度。一个基因可以在3 '端表达多种不同的同种异构体，这种现象被称为选择性切割和聚腺苷酸化，或APA。以往的研究表明，大多数真核生物基因都显示APA， APA谱在不同的生理和病理条件下发生变化。然而，APA在细胞中的调节机制尚不清楚。在这里，我们使用基因敲低和高通量测序来研究APA是如何被负责切割和聚腺苷化的机制中的因子以及在剪接中起重要作用的因子所调节的。我们在最后一个外显子中发现了几个在APA中起重要作用的因子，包括CFI-25/68、PABPN1、PABPC1、Fip1和Pcf11。我们还阐明了内含子和转录起始位点附近的切割和聚腺苷化事件是如何被调节的。我们发现了一组APA事件，这些事件受到核心因子以及细胞分化和发育的高度调节。我们提出了一个APA代码，其中在给定的细胞环境中，APA事件受到许多参数的调节，包括转录起始位点的相对位置，剪接环境，竞争pAs之间的距离，周围的顺式元件以及核心切割和聚腺苷化因子的浓度。
Alternative cleavage and polyadenylation (APA) results in mRNA isoforms containing different 3’ untranslated regions (3’UTRs) and/or coding sequences. How core cleavage/polyadenylation (C/P) factors regulate APA is not well understood. Using siRNA knockdown coupled with deep sequencing, we found that several C/P factors can play significant roles in 3’UTR-APA. Whereas Pcf11 and Fip1 enhance usage of proximal poly(A) sites (pAs), CFI-25/68, PABPN1 and PABPC1 promote usage of distal pAs. Strong cis element biases were found for pAs regulated by CFI-25/68 or Fip1, and the distance between pAs plays an important role in APA regulation. In addition, intronic pAs are substantially regulated by splicing factors, with U1 mostly inhibiting C/P events in introns near the 5’ end of gene and U2 suppressing those in introns with features for efficient splicing. Furthermore, PABPN1 inhibits expression of transcripts with pAs near the transcription start site (TSS), a property possibly related to its role in RNA degradation. Finally, we found that groups of APA events regulated by C/P factors are also modulated in cell differentiation and development with distinct trends. Together, our results support an APA code where an APA event in a given cellular context is regulated by a number of parameters, including relative location to the TSS, splicing context, distance between competing pAs, surrounding cis elements and concentrations of core C/P factors. A gene can express multiple isoforms varying in the 3’ end, a phenomenon called alternative cleavage and polyadenylation, or APA. Previous studies have indicated that most eukaryotic genes display APA and the APA profile changes under different physiological and pathological conditions. However, how APA is regulated in the cell is unclear. Here using gene knockdown and high throughput sequencing we examine how APA is regulated by factors in the machinery responsible for cleavage and polyadenylation as well as factors that play essential roles in splicing. We identify several factors that play significant roles in APA in the last exon, including CFI-25/68, PABPN1, PABPC1, Fip1 and Pcf11. We also elucidate how cleavage and polyadenylation events are regulated in introns and near the transcription start site. We uncover a group of APA events that are highly regulated by core factors as well as in cell differentiation and development. We present an APA code where an APA event in a given cellular context is regulated by a number of parameters, including relative location to the transcription start site, splicing context, distance between competing pAs, surrounding cis elements and concentrations of core cleavage and polyadenylation factors.
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