Oliver Stefan L, Sommer Marvin, Zerboni Leigh, Rajamani Jaya, Grose Charles, Arvin Ann M
Department of Pediatrics, Stanford University School of Medicine, Stanford, California 94305, USA.
J Virol. 2009 Aug;83(15):7495-506. doi: 10.1128/JVI.00400-09. Epub 2009 May 27.
Glycoprotein B (gB), the most conserved protein in the family Herpesviridae, is essential for the fusion of viral and cellular membranes. Information about varicella-zoster virus (VZV) gB is limited, but homology modeling showed that the structure of VZV gB was similar to that of herpes simplex virus (HSV) gB, including the putative fusion loops. In contrast to HSV gB, VZV gB had a furin recognition motif ([R]-X-[KR]-R-|-X, where | indicates the position at which the polypeptide is cleaved) at residues 491 to 494, thought to be required for gB cleavage into two polypeptides. To investigate their contribution, the putative primary fusion loop or the furin recognition motif was mutated in expression constructs and in the context of the VZV genome. Substitutions in the primary loop, W180G and Y185G, plus the deletion mutation Delta491RSRR494 and point mutation 491GSGG494 in the furin recognition motif did not affect gB expression or cellular localization in transfected cells. Infectious VZV was recovered from parental Oka (pOka)-bacterial artificial chromosomes that had either the Delta491RSRR494 or 491GSGG494 mutation but not the point mutations W180G and Y185G, demonstrating that residues in the primary loop of gB were essential but gB cleavage was not required for VZV replication in vitro. Virion morphology, protein localization, plaque size, and replication were unaffected for the pOka-gBDelta491RSRR494 or pOka-gB491GSGG494 virus compared to pOka in vitro. However, deletion of the furin recognition motif caused attenuation of VZV replication in human skin xenografts in vivo. This is the first evidence that cleavage of a herpesvirus fusion protein contributes to viral pathogenesis in vivo, as seen for fusion proteins in other virus families.
糖蛋白B（gB）是疱疹病毒科中最保守的蛋白质，对于病毒膜与细胞膜的融合至关重要。关于水痘带状疱疹病毒（VZV）gB的信息有限，但同源建模显示VZV gB的结构与单纯疱疹病毒（HSV）gB相似，包括推定的融合环。与HSV gB不同，VZV gB在第491至494位残基处有一个弗林蛋白酶识别基序（[R]-X-[KR]-R-|-X，其中|表示多肽被切割的位置），被认为是gB切割成两条多肽所必需的。为了研究它们的作用，在表达构建体和VZV基因组背景下对推定的主要融合环或弗林蛋白酶识别基序进行了突变。主要环中的取代W180G和Y185G，以及弗林蛋白酶识别基序中的缺失突变Delta491RSRR494和点突变491GSGG494，并不影响转染细胞中gB的表达或细胞定位。从具有Delta491RSRR494或491GSGG494突变但没有点突变W180G和Y185G的亲本Oka（pOka）-细菌人工染色体中回收了感染性VZV，这表明gB主要环中的残基是必不可少的，但gB切割对于VZV体外复制不是必需的。与体外的pOka相比，pOka-gBDelta491RSRR494或pOka-gB491GSGG494病毒的病毒体形态、蛋白质定位、蚀斑大小和复制均未受影响。然而，弗林蛋白酶识别基序的缺失导致VZV在体内人皮肤异种移植中的复制减弱。这是首次有证据表明疱疹病毒融合蛋白的切割有助于病毒在体内的发病机制，这与其他病毒家族中的融合蛋白情况相同。
