Ubiquitylation of the acetyltransferase MOF in Drosophila melanogaster.
Ubiquitylation of the acetyltransferase MOF in Drosophila melanogaster.
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DOI：
10.1371/journal.pone.0177408
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发表时间：
2017
期刊：
影响因子：
3.7
通讯作者：
Becker PB
中科院分区：
文献类型：
--
作者：
Schunter S;Villa R;Flynn V;Heidelberger JB;Classen AK;Beli P;Becker PB
核乙酰转移酶 MOF（哺乳动物中的 KAT8）是至少两个参与转录调节的多组分复合物的亚基。在“非特异性致死”（NSL）类型复合物的背景下，它控制许多核管家基因和线粒体基因的转录起始。虽然这种功能在后生动物中是保守的，但 MOF 在果蝇中具有剂量补偿方面的附加、特定功能。作为男性特异性致死剂量补偿复合物 (MSL-DCC) 的一个亚基，它通过乙酰化组蛋白 H4 来使单个男性 X 染色体的转录输出加倍。适当的剂量补偿需要精细调节 MSL-DCC 的水平以及调节复合物之间 MOF 的适当分布。 DCC 的形成量直接取决于男性特异性 MSL2 的水平，MSL2 负责协调 DCC 的组装，包括 MOF 的招募。我们之前发现 MSL2 是一种 E3 连接酶，可泛素化大多数 MSL 蛋白，包括 MOF，这表明泛素化可能有助于 MOF 整体水平和折叠状态及其在复杂实体之间的分配的质量控制。现在，我们使用质谱法绘制了 MOF 中在体外被 MSL2 泛素化的赖氨酸图谱，并鉴定了 MOF 在雄性和雌性细胞中的体内泛素化位点。由于其他性别无关的泛素化事件占主导地位，因此无法追踪体内 MSL2 特异性泛素化，并且可以想象，这种泛素化可能是罕见的或短暂的。通过表达适当突变的 MOF 衍生物，我们通过监测培养细胞中 DCC 的形成和 X 染色体靶向，以及果蝇中雄性特异性致死 mof2 等位基因的遗传互补，评估了泛素化赖氨酸对于剂量补偿的重要性。我们的研究对MOF泛素化进行了全面的分析，为未来的研究提供参考。
The nuclear acetyltransferase MOF (KAT8 in mammals) is a subunit of at least two multi-component complexes involved in transcription regulation. In the context of complexes of the ‘Non-Specific-Lethal’ (NSL) type it controls transcription initiation of many nuclear housekeeping genes and of mitochondrial genes. While this function is conserved in metazoans, MOF has an additional, specific function in Drosophila in the context of dosage compensation. As a subunit of the male-specific-lethal dosage compensation complex (MSL-DCC) it contributes to the doubling of transcription output from the single male X chromosome by acetylating histone H4. Proper dosage compensation requires finely tuned levels of MSL-DCC and an appropriate distribution of MOF between the regulatory complexes. The amounts of DCC formed depends directly on the levels of the male-specific MSL2, which orchestrates the assembly of the DCC, including MOF recruitment. We found earlier that MSL2 is an E3 ligase that ubiquitylates most MSL proteins, including MOF, suggesting that ubiquitylation may contribute to a quality control of MOF’s overall levels and folding state as well as its partitioning between the complex entities. We now used mass spectrometry to map the lysines in MOF that are ubiquitylated by MSL2 in vitro and identified in vivo ubiquitylation sites of MOF in male and female cells. MSL2-specific ubiquitylation in vivo could not be traced due to the dominance of other, sex-independent ubiquitylation events and conceivably may be rare or transient. Expressing appropriately mutated MOF derivatives we assessed the importance of the ubiquitylated lysines for dosage compensation by monitoring DCC formation and X chromosome targeting in cultured cells, and by genetic complementation of the male-specific-lethal mof2 allele in flies. Our study provides a comprehensive analysis of MOF ubiquitylation as a reference for future studies.