Chromatin Organization and RNA silencing

染色质组织和 RNA 沉默

• 批准号: 8148677
• 负责人: Elissa Lei
• 金额: $ 32.93万
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8148677
• 关键词: Chromatin; RNA Interference

In order to determine if these Argonaute genes are required for heterochromatin formation, we utilized transcriptional reporters and ChIP of HP1 from somatic head tissue to monitor the heterochromatic state of piRNA clusters, which generate both endo-siRNAs and the bulk of piRNAs. Surprisingly, we find that mutation of AGO2 or piwi increases silencing at piRNA clusters corresponding to an increase of HP1 association. Furthermore, loss of piRNA production from a single piRNA cluster results in genome-wide redistribution of HP1 and reduction of silencing at a distant heterochromatic site suggesting indirect effects on HP1 localization. Taken together, these results indicate that heterochromatin forms independently of endo-siRNA and piRNA pathways in Drosophila somatic tissues. Our results indicate that heterochromatin can form independently of these two small RNA silencing pathways and appears to be achieved by alternate mechanisms in Drosophila and perhaps other metazoans. To further elucidate the role of Argonaute proteins in the higher order chromatin function, we performed ChIP followed by Illumina sequencing on an ovarian somatic follicle cell line. Our analysis revealed no association of Piwi with euchromatin but enrichment at highly repetitive sequences. In contrast, AGO2 localizes primarily with euchromatic sites throughout the genome.

为了确定异染色质形成是否需要这些ARGONAUTE基因，我们利用了转录报告基因和HP1的芯片从体型头组织监测PIRNA簇的异质状状态，这同时会产生内部siRNAS和PIRNA的大部分。 令人惊讶的是，我们发现AGO2或PIWI的突变会增加与HP1缔合增加相对应的PIRNA簇的沉默。 此外，单个piRNA簇因piRNA产生的损失导致HP1的全基因组重新分布和在远处异质体部位减少沉默，这表明对HP1定位的间接影响。 综上所述，这些结果表明，异染色质在果蝇体细胞组织中独立于内核和piRNA途径而形成。 我们的结果表明，异染色质可以独立于这两个小的RNA沉默途径形成，并且似乎是通过果蝇和其他后生动物中的其他机制来实现的。 为了进一步阐明Argonaute蛋白在高阶染色质功能中的作用，我们进行了芯片，然后在卵巢体细胞卵泡细胞系上进行了Illumina测序。 我们的分析表明，Piwi与斑塑素没有关联，而是在高度重复的序列上富集。 相比之下，AGO2主要与整个基因组中的正式位点定位。