Interaction of Bombyx mori nucleopolyhedrovirus BRO-A and host cell protein laminin
Interaction of Bombyx mori nucleopolyhedrovirus BRO-A and host cell protein laminin
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DOI：
10.1007/s00705-002-0902-7
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发表时间：
2003-01-01
影响因子：
2.7
通讯作者：
Zemskov, EA
中科院分区：
文献类型：
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作者：
Kang, WK;Imai, N;Zemskov, EA
家蚕核多角体病毒 (BmNPV) 含有 5 个杆状病毒重复 ORF (bro) 基因，所有这些基因均表达为延迟早期基因。我们最近报道了 BmNPV BRO 蛋白，特别是 BRO-A 和 BRO-C，具有核酸结合活性，并参与受感染细胞核中的核小体结构。为了进一步了解 bro-a 基因的功能，我们使用酵母双杂交系统寻找与 BmNPV BRO-A 相互作用的因子。发现从模拟感染细胞的 cDNA 文库中获得的 15 个克隆和从感染后 2 小时 (p.i.) 制备的文库中获得的一个克隆包含一个不同的基因，该基因被鉴定为果蝇层粘连蛋白 beta1 的家蚕同源物 (bLaminin)。体外下拉实验证明了 BRO-A 和 bLaminin N 端区域之间的直接相互作用。使用 BmN 细胞提取物和抗层粘连蛋白抗体进行的进一步 Pull-down 测定也显示了两种蛋白质的相互作用。此外，从注射后12小时的cDNA文库中还获得了另外两个克隆。并被发现编码 BRO-A 本身，表明 BRO-A 形成寡聚物。综上所述，我们推测 BRO-A 可能充当层粘连蛋白结合蛋白。
The Bombyx mori nucleopolyhedrovirus (BmNPV) contains five baculovirus repeated ORF (bro) genes, all of which are expressed as delayed early genes. We have recently reported that BmNPV BRO proteins, specially BRO-A and BRO-C, contain a nucleic acid binding activity and are involved in nucleosome structures in nuclei of infected cells. To further understand the function of bro-a gene, we looked for factors interacting with BmNPV BRO-A using the yeast two-hybrid system. Fifteen clones obtained from a cDNA library of mock-infected cells and one from a library prepared at 2 h postinfection (p.i.) were found to comprise one distinct gene, which was identified as the Bombyx homolog (bLaminin) of Drosophila laminin beta1. A direct interaction between BRO-A and N-terminal region of bLaminin was demonstrated by in vitro pull-down experiments. Further pull-down assays using BmN cell extracts and antilaminin antibodies also showed interaction of both proteins. In addition, two more clones were obtained from cDNA library of 12 h p.i. and were found to encode BRO-A itself, indicating that BRO-A forms an oligomer. Taken together, we propose that BRO-A may function as a laminin binding protein.