Mijnes J D, van der Horst L M, van Anken E, Horzinek M C, Rottier P J, de Groot R J
Virology Unit, Department of Infectious Diseases, Utrecht University, The Netherlands.
J Virol. 1996 Aug;70(8):5466-75. doi: 10.1128/JVI.70.8.5466-5475.1996.
The biosynthesis of glycoproteins E and I of feline herpesvirus was studied by using the vaccinia virus vTF7-3 expression system. gE and gI were synthesized as N-glycosylated, endoglycosidase H (EndoH)-sensitive precursors with Mrs of 83,000 and 67,000, respectively. When coexpressed, gE and gI formed sodium dodecyl sulfate-sensitive hetero-oligomeric complexes that were readily transported from the endoplasmic reticulum (ER). Concomitantly, the glycoproteins acquired extensive posttranslational modifications, including O glycosylation, leading to an increase in their apparent molecular weights to 95,000 and 80,000 to 100,000 for gE and gI, respectively. In the absence of gE, most gI remained EndoH sensitive. Only a minor population became EndoH resistant, but these molecules were processed aberrantly as indicated by their Mrs (100,000 to 120,000). By immunofluorescence microscopy, gI was detected primarily in the ER but also at the plasma membrane. gE, when expressed by itself, remained EndoH sensitive and was found only in the ER and the nuclear envelope. These results were corroborated by studying the biosynthesis of gE in feline herpesvirus (FHV)-infected cells. In cells infected with wild-type FHV, gE acquired the same co- and posttranslational modifications as during vTF7-3-driven expression. However, an FHV mutant lacking gI failed to produce mature gE. We conclude that gE is retained in the ER, presumably by associating with molecular chaperones, and becomes transport competent only when in a complex with gI.
利用痘苗病毒vTF7-3表达系统研究了猫疱疹病毒糖蛋白E和I的生物合成。gE和gI作为N-糖基化的、对内切糖苷酶H(EndoH)敏感的前体进行合成,其分子量分别为83,000和67,000。共表达时,gE和gI形成对十二烷基硫酸钠敏感的异源寡聚复合物,这些复合物很容易从内质网(ER)转运出去。同时,这些糖蛋白进行了广泛的翻译后修饰,包括O-糖基化,导致它们的表观分子量分别增加到gE为95,000,gI为80,000至100,000。在没有gE的情况下,大多数gI仍对EndoH敏感。只有一小部分变为对EndoH有抗性,但这些分子的加工异常,这从它们的分子量(100,000至120,000)可以看出。通过免疫荧光显微镜观察,gI主要在内质网中检测到,但也在质膜上检测到。gE单独表达时,仍对EndoH敏感,仅在内质网和核膜中发现。通过研究猫疱疹病毒(FHV)感染细胞中gE的生物合成,证实了这些结果。在感染野生型FHV的细胞中,gE获得了与vTF7-3驱动表达期间相同的共翻译和翻译后修饰。然而,缺乏gI的FHV突变体未能产生成熟的gE。我们得出结论,gE保留在内质网中,推测是通过与分子伴侣结合,并且只有在与gI形成复合物时才具有转运能力。