Chen Jiaying, Liu Na, Qi Hongying, Neuenkirchen Nils, Huang Yuedong, Lin Haifan
Yale Stem Cell Center, 10 Amistad St., Room 237E, New Haven, CT 06511, USA.
Department of Genetics, 333 Cedar St., New Haven, CT 06511, USA.
Nucleic Acids Res. 2025 Jan 7;53(1). doi: 10.1093/nar/gkae1160.
Alternative transcription initiation, which refers to the transcription of a gene from different transcription start sites (TSSs), is prevalent across metazoans and has important biological functions. Although transcriptional regulation has been extensively studied, the mechanism that selects one TSS over others within a gene remains elusive. Using the Cap Analysis of Gene Expression sequencing (CAGE-seq) method, we discovered that Piwi, an RNA-binding protein, regulates TSS usage in at least 87 genes. In piwi-deficient Drosophila ovaries, these genes displayed significantly altered TSS usage (ATU). The regulation of TSS usage occurred in both germline and somatic cells in ovaries, as well as in cultured ovarian somatic cells (OSCs). Correspondingly, RNA Polymerase II (Pol II) initiation and elongation at the TSSs of ATU genes were affected in germline-piwi-knockdown ovaries and piwi-knockdown OSCs. Furthermore, we identified a Facilitates Chromatin Transcription (FACT) complex component, Ssrp, that is essential for mRNA elongation, as a novel interactor of Piwi in the nucleus. Temporally controlled knockdown of ssrp affected TSS usage in ATU genes, whereas overexpression of ssrp partially rescued the TSS usage of ATU genes in piwi mutant ovaries. Thus, Piwi may interact with Ssrp to regulate TSS usage in Drosophila ovaries by affecting Pol II initiation and elongation.
可变转录起始是指基因从不同的转录起始位点（TSS）进行转录，这种现象在后生动物中普遍存在，并具有重要的生物学功能。尽管转录调控已得到广泛研究，但在一个基因内选择一个TSS而非其他TSS的机制仍不清楚。我们使用基因表达测序的帽分析（CAGE-seq）方法发现，RNA结合蛋白Piwi可调控至少87个基因的TSS使用情况。在缺乏Piwi的果蝇卵巢中，这些基因的TSS使用情况发生了显著改变（ATU）。TSS使用情况的调控发生在卵巢的生殖细胞和体细胞中，以及培养的卵巢体细胞（OSC）中。相应地，在生殖系Piwi基因敲低的卵巢和Piwi基因敲低的OSC中，ATU基因TSS处的RNA聚合酶II（Pol II）起始和延伸受到影响。此外，我们鉴定出一种促进染色质转录（FACT）复合体成分Ssrp，它对mRNA延伸至关重要，是Piwi在细胞核中的一种新的相互作用蛋白。对Ssrp进行时间控制的敲低会影响ATU基因的TSS使用情况，而Ssrp的过表达可部分挽救Piwi突变卵巢中ATU基因的TSS使用情况。因此，Piwi可能通过影响Pol II起始和延伸与Ssrp相互作用，从而调控果蝇卵巢中的TSS使用情况。
