The Drosophila maternal-effect gene abnormal oocyte (ao) does not repress histone gene expression

Risa Takenaka, Sierra M Simmerman, Casey A Schmidt, Eric H Albanese, Leila E Rieder, Harmit Singh Malik
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Abstract

The abnormal oocyte (ao) gene of Drosophila melanogaster is a maternal-effect lethal gene previously identified as encoding a transcriptional regulator of core histones. However, background genetic mutations in existing ao mutant strains could compromise their utility in manipulating histone levels. To distinguish the true ao pheno-type from background effects, we created two new ao reagents: a CRISPR/Cas9-mediated knockout of the ao allele for genetic and molecular analyses and an epitope-tagged ao allele for cytological experiments. Using these rea-gents, we confirm previous findings that ao exhibits maternal-effect lethality, which can be rescued by either a decrease in the histone gene copy number or by Y chromosome heterochromatin. We also confirm that the Ao protein localizes to the histone locus bodies in ovaries. Our data also suggest that ao genetically interacts with the histone genes and heterochromatin, as previously suggested. However, contrary to prior findings, we find that ao does not repress core histone transcript levels. Thus, the molecular basis for ao-associated maternal-effect lethality remains unknown.
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果蝇的母性效应基因异常卵母细胞(ao)不会抑制组蛋白基因的表达
黑腹果蝇的异常卵母细胞(ao)基因是一种母系效应致死基因,以前曾被鉴定为编码核心组蛋白的转录调节因子。然而,现有ao突变株系中的背景基因突变可能会影响其在组蛋白水平操作中的效用。为了区分真正的 ao 表型和背景效应,我们创建了两种新的 ao 试剂:一种是 CRISPR/Cas9 介导的 ao 等位基因敲除,用于遗传和分子分析;另一种是表位标记的 ao 等位基因,用于细胞学实验。利用这些等位基因,我们证实了之前的发现,即 ao 表现出母性效应致死性,而这种致死性可以通过组蛋白基因拷贝数的减少或 Y 染色体异染色质来挽救。我们还证实,Ao 蛋白定位于卵巢中的组蛋白基因座体。我们的数据还表明,鳌与组蛋白基因和异染色质之间存在基因相互作用,这与之前的研究结果一致。然而,与之前的发现相反,我们发现鳌并不抑制核心组蛋白转录本的水平。因此,ao 相关母性效应致死的分子基础仍然未知。
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