Institute of Cell Biology, University of Bern, Bern, Switzerland.
Graduate School for Cellular and Biomedical Sciences, University of Bern, Bern, Switzerland.
EMBO J. 2022 Nov 17;41(22):e111839. doi: 10.15252/embj.2022111839. Epub 2022 Oct 11.
Small RNAs mediate the silencing of transposable elements and other genomic loci, increasing nucleosome density and preventing undesirable gene expression. The unicellular ciliate Paramecium is a model to study dynamic genome organization in eukaryotic cells, given its unique feature of nuclear dimorphism. Here, the formation of the somatic macronucleus during sexual reproduction requires eliminating thousands of transposon remnants (IESs) and transposable elements scattered throughout the germline micronuclear genome. The elimination process is guided by Piwi-associated small RNAs and leads to precise cleavage at IES boundaries. Here we show that IES recognition and precise excision are facilitated by recruiting ISWI1, a Paramecium homolog of the chromatin remodeler ISWI. ISWI1 knockdown substantially inhibits DNA elimination, quantitatively similar to development-specific sRNA gene knockdowns but with much greater aberrant IES excision at alternative boundaries. We also identify key development-specific sRNA biogenesis and transport proteins, Ptiwi01 and Ptiwi09, as ISWI1 cofactors in our co-immunoprecipitation studies. Nucleosome profiling indicates that increased nucleosome density correlates with the requirement for ISWI1 and other proteins necessary for IES excision. We propose that chromatin remodeling together with small RNAs is essential for efficient and precise DNA elimination in Paramecium.
小 RNA 介导转座元件和其他基因组位点的沉默,增加核小体密度,防止不期望的基因表达。单细胞纤毛虫草履虫是研究真核细胞动态基因组组织的模型,因为其核二态性的独特特征。在这里,有性生殖过程中体细胞大核的形成需要消除散布在生殖系微核基因组中的数千个转座子残余物(IESs)和转座元件。消除过程由 Piwi 相关小 RNA 指导,并导致 IES 边界的精确切割。在这里,我们表明 IES 的识别和精确切除是通过招募 Paramecium 同源物染色质重塑酶 ISWI 的 ISWI1 来促进的。ISWI1 的敲低会极大地抑制 DNA 的消除,与特定发育阶段的小 RNA 基因敲低的效果非常相似,但在替代边界处会出现更多异常的 IES 切除。我们还鉴定了关键的发育特异性小 RNA 生物发生和转运蛋白 Ptiwi01 和 Ptiwi09,它们是我们共免疫沉淀研究中 ISWI1 的共因子。核小体谱分析表明,核小体密度的增加与 IES 切除所需的 ISWI1 和其他蛋白有关。我们提出,染色质重塑与小 RNA 一起对于草履虫中高效和精确的 DNA 消除是必不可少的。