Evidence for Nucleic Acid Binding Ability and Nucleosome Association of Bombyx mori Nucleopolyhedrovirus BRO Proteins
Evidence for Nucleic Acid Binding Ability and Nucleosome Association of Bombyx mori Nucleopolyhedrovirus BRO Proteins
复制标题
DOI：
10.1128/jvi.74.15.6784-6789.2000
复制
发表时间：
2000-08
影响因子：
5.4
通讯作者：
E. Zemskov;W. Kang;S. Maeda
中科院分区：
文献类型：
--
作者：
E. Zemskov;W. Kang;S. Maeda
家蚕核型多角体病毒（BmNPV）基因组包含5个bro基因家族的相关成员，它们都在感染的BmN细胞中活跃表达。虽然它们的功能是未知的，但它们的氨基酸序列含有在所有已知的病毒和原核单链DNA结合蛋白中发现的基序。为了确定它们是否与核酸结合，我们使用组蛋白提取方案分离了BmNPV感染的BmN细胞的细胞核。我们使用抗BRO抗体检测组蛋白H1组分中的BRO-A、BRO-C和BRO-D。微球菌核酸酶处理释放这些BRO蛋白质的染色质部分，表明它们参与核小体结构。色谱分离表明BRO-A和/或BRO-C与核心组蛋白相互作用。BRO-A部分序列的表达证明N端80个氨基酸残基是DNA结合活性所需的。我们还表明，BmNPV BRO蛋白经历磷酸化和泛素化，然后蛋白酶体降解，这可能解释了它们在细胞质以及细胞核中的分布。我们建议，BRO-A和BRO-C可能作为DNA结合蛋白，影响宿主DNA复制和/或转录。
ABSTRACT The Bombyx mori nucleopolyhedrovirus (BmNPV) genome contains five related members of the bro gene family, all of which are actively expressed in infected BmN cells. Although their functions are unknown, their amino acid sequences contain a motif found in all known viral and prokaryotic single-stranded DNA binding proteins. To determine if they bind to nucleic acids, we fractionated the nuclei of BmNPV-infected BmN cells using a histone extraction protocol. We detected BRO-A, BRO-C, and BRO-D in the histone H1 fraction using anti-BRO antibodies. Micrococcal nuclease treatment released these BRO proteins from the chromatin fraction, suggesting their involvement in nucleosome structures. Chromatographic fractionation showed that BRO-A and/or BRO-C interacted with core histones. Expression of partial sequences of BRO-A proved that the N-terminal 80 amino acid residues were required for DNA binding activity. We also demonstrated that BmNPV BRO proteins underwent phosphorylation and ubiquitination followed by proteasome degradation, which may explain their distribution in the cytoplasm as well as the nucleus. We propose that BRO-A and BRO-C may function as DNA binding proteins that influence host DNA replication and/or transcription.