Wu E, Fernandez J, Fleck S K, Huang S, Nemerow G R
Department of Immunology, The Scripps Research Institute, La Jolla, California, 92037
Virology. 2001 Jan 5;279(1):78-89. doi: 10.1006/viro.2000.0703.
The ocular tropism of adenovirus type 37 (Ad37) does not correlate with the wide distribution of the 46-kDa coxsackievirus and adenovirus receptor (CAR), the major receptor for most adenovirus serotypes. We previously found that Ad37 infects and binds well to conjunctival cells (Chang C), but poorly to lung epithelial (A549) cells that express CAR and hypothesized that this serotype uses a distinct receptor that is selectively expressed on conjunctival cells. To test this, we produced particles of a fiber-deleted Ad5 vector containing the Ad37 fiber protein. The "pseudotyped" vector infected Chang C cells better than A549 cells using a CAR-independent pathway. Ad37 binding was calcium-dependent and was abolished by protease digestion of cell surface proteins. Using a virus overlay protein blot assay (VOPBA), we detected calcium-dependent Ad37 binding to 50- and 60-kDa membrane proteins on permissive Chang C cells. In contrast, calcium-dependent binding was detected with only the 60-kDa protein on nonpermissive A549 cells. Ad19p, a closely related serotype that failed to bind to conjunctival cells, recognized the 60-kDa, but not the 50-kDa, protein. Ad37 has been reported to use sialic acid instead of CAR as a cell receptor on A549 cells. Pretreatment of Chang C cells with neuraminidase abolished Ad37 binding to only the 60-kDa protein, suggesting that sialic acid mediates Ad37 binding to the 60-kDa protein. The pseudotyped Ad37 vector was also able to infect neuraminidase-treated Chang C cells. Thus, subgroup D adenoviral binding to the 50-kDa protein is calcium-dependent and cell type- and serotype-specific, whereas binding to the 60-kDa protein is not necessary for infection of conjunctival cells. Together, these data suggest that the 50-kDa protein is the major receptor for Ad37 on conjunctival cells.
37型腺病毒（Ad37）的眼嗜性与46 kDa柯萨奇病毒和腺病毒受体（CAR）的广泛分布不相关，CAR是大多数腺病毒血清型的主要受体。我们之前发现Ad37能很好地感染并结合结膜细胞（Chang C），但对表达CAR的肺上皮（A549）细胞的结合能力较差，并推测该血清型使用一种在结膜细胞上选择性表达的独特受体。为了验证这一点，我们制备了含有Ad37纤维蛋白的缺失纤维的Ad５载体颗粒。这种“假型”载体通过一种不依赖CAR的途径，对Chang C细胞的感染能力优于A549细胞。Ad37的结合依赖于钙，并且通过蛋白酶消化细胞表面蛋白可消除这种结合。使用病毒覆盖蛋白印迹分析（VOPBA），我们检测到Ad37与允许性Chang C细胞上50 kDa和60 kDa的膜蛋白存在钙依赖性结合。相比之下，在非允许性A549细胞上仅检测到与６０kDa蛋白的钙依赖性结合。Ad19p是一种密切相关的血清型，它不能与结膜细胞结合，能识别60 kDa的蛋白，但不能识别50 kDa的蛋白。据报道，Ad37在A549细胞上使用唾液酸而非CAR作为细胞受体。用神经氨酸酶预处理Chang C细胞仅消除了Ad37与60 kDa蛋白的结合，这表明唾液酸介导了Ad37与60 kDa蛋白的结合。这种假型Ad37载体也能够感染经神经氨酸酶处理的Chang C细胞。因此，D亚组腺病毒与50 kDa蛋白的结合是钙依赖性的，并且具有细胞类型和血清型特异性，而与60 kDa蛋白的结合对于结膜细胞的感染并非必需。总之，这些数据表明50 kDa蛋白是Ad37在结膜细胞上的主要受体。
