Kobayashi T, Shoya Y, Koda T, Takashima I, Lai P K, Ikuta K, Kakinuma M, Kishi M
Institute of Immunological Science, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo, Japan.
Virology. 1998 Mar 30;243(1):188-97. doi: 10.1006/viro.1998.9049.
The Borna disease virus (BDV) replicates in the nucleus. The viral p40 protein (N), which is found abundantly in the nucleus in BDV-infected cells, may play an important role in virus replication. To analyze the amino acid residues involved in the nuclear targeting of BDV N, a series of eukaryotic expression plasmids encoding deletion mutants of N was constructed and transfected into COS-7 cells. In indirect immunofluorescence assays with a rabbit anti-BDV N antiserum, wild-type N was located in the nucleus of transfected cells in the absence of other viral constituents. In contrast, mutants lacking the 13 NH2-terminal amino acid residues 1MPPKRRLVDDADA13 in common gave a cytoplasmic localization pattern. Similarly, a mutant with substitution of 4KRR6 by 4NSG6 was retained in the cytoplasm. Furthermore, a nonapeptide, 3PKRRLVDDA11, derived from the NH2-terminal region of N conferred nuclear targeting activity to beta-galactosidase, which normally resides in the cytoplasm. Thus, we have identified the nuclear targeting signal of the BDV N and narrowed it to the NH2-terminal region where 4KRR6 basic amino acid residues are located.
博尔纳病病毒（BDV）在细胞核中复制。病毒的p40蛋白（N）在BDV感染的细胞的细胞核中大量存在，可能在病毒复制中发挥重要作用。为了分析参与BDV N核靶向的氨基酸残基，构建了一系列编码N缺失突变体的真核表达质粒，并将其转染到COS-7细胞中。在用兔抗BDV N抗血清进行的间接免疫荧光试验中，在没有其他病毒成分的情况下，野生型N位于转染细胞的细胞核中。相反，共同缺失13个氨基末端氨基酸残基1MPPKRRLVDDADA13的突变体呈现出细胞质定位模式。同样，将4KRR6替换为4NSG6的突变体保留在细胞质中。此外，源自N氨基末端区域的九肽3PKRRLVDDA11赋予通常位于细胞质中的β-半乳糖苷酶核靶向活性。因此，我们已经鉴定出BDV N的核靶向信号，并将其缩小到4KRR6碱性氨基酸残基所在的氨基末端区域。
