Ciarlet Max, Crawford Sue E, Cheng Elly, Blutt Sarah E, Rice Daren A, Bergelson Jeffrey M, Estes Mary K
Department of Molecular Virology and Microbiology, Baylor College of Medicine, Houston, Texas 77030, USA.
J Virol. 2002 Feb;76(3):1109-23. doi: 10.1128/jvi.76.3.1109-1123.2002.
In an attempt to identify the rotavirus receptor, we tested 46 cell lines of different species and tissue origins for susceptibility to infection by three N-acetyl-neuraminic (sialic) acid (SA)-dependent and five SA-independent rotavirus strains. Susceptibility to SA-dependent or SA-independent rotavirus infection varied depending on the cell line tested and the multiplicity of infection (MOI) used. Cells of renal or intestinal origin and transformed cell lines derived from breast, stomach, bone, or lung were all susceptible to rotavirus infection, indicating a wider host tissue range than previously appreciated. Chinese hamster ovary (CHO), baby hamster kidney (BHK-21), guinea pig colon (GPC-16), rat small intestine (Rie1), and mouse duodenum (MODE-K) cells were found to support only limited rotavirus replication even at MOIs of 100 or 500, but delivery of rotavirus particles into the cytoplasm by lipofection resulted in efficient rotavirus replication. The rotavirus cell attachment protein, the outer capsid spike protein VP4, contains the sequence GDE(A) recognized by the VLA-2 (alpha2beta1) integrin, and to test if VLA-2 is involved in rotavirus attachment and entry, we measured infection in CHO cells that lack VLA-2 and CHO cells transfected with the human alpha2 subunit (CHOalpha2) or with both the human alpha2 and beta1 subunits (CHOalpha2beta1) of VLA-2. Infection by SA-dependent or SA-independent rotavirus strains was 2- to 10-fold more productive in VLA-2-expressing CHO cells than in parental CHO cells, and the increased susceptibility to infection was blocked with anti-VLA-2 antibody. However, the levels of binding of rotavirus to CHO, CHOalpha2, and CHOalpha2beta1 cells were equivalent and were not increased over binding to susceptible monkey kidney (MA104) cells or human colonic adenocarcinoma (Caco-2, HT-29, and T-84) cells, and binding was not blocked by antibody to the human alpha2 subunit. Although the VLA-2 integrin promotes rotavirus infection in CHO cells, it is clear that the VLA-2 integrin alone is not responsible for rotavirus cell attachment and entry. Therefore, VLA-2 is not involved in the initial attachment of rotavirus to cells but may play a role at a postattachment level.
为了鉴定轮状病毒受体，我们检测了46种不同物种和组织来源的细胞系对3种依赖N - 乙酰神经氨酸（唾液酸，SA）的和5种不依赖SA的轮状病毒株感染的易感性。对依赖SA或不依赖SA的轮状病毒感染的易感性因所检测的细胞系和所用的感染复数（MOI）而异。肾或肠来源的细胞以及源自乳腺、胃、骨或肺的转化细胞系均对轮状病毒感染敏感，这表明宿主组织范围比之前认为的更广。发现中国仓鼠卵巢（CHO）细胞、幼仓鼠肾（BHK - 21）细胞、豚鼠结肠（GPC - 16）细胞、大鼠小肠（Rie1）细胞和小鼠十二指肠（MODE - K）细胞即使在MOI为100或500时也仅支持有限的轮状病毒复制，但通过脂质体转染将轮状病毒颗粒递送至细胞质中可导致高效的轮状病毒复制。轮状病毒细胞附着蛋白，即外衣壳刺突蛋白VP4，含有被VLA - 2（α2β1）整合素识别的序列GDE(A)，为了检测VLA - 2是否参与轮状病毒的附着和进入，我们检测了缺乏VLA - 2的CHO细胞以及转染了人α2亚基（CHOα2）或同时转染了人α2和β1亚基（CHOα2β1）的VLA - 2的CHO细胞中的感染情况。与亲本CHO细胞相比，表达VLA - 2的CHO细胞中依赖SA或不依赖SA的轮状病毒株的感染效率提高了2至10倍，并且抗VLA - 2抗体可阻断感染易感性的增加。然而，轮状病毒与CHO、CHOα2和CHOα2β1细胞的结合水平相当，与易感的猴肾（MA104）细胞或人结肠腺癌（Caco - 2、HT - 29和T - 84）细胞的结合相比并未增加，并且人α2亚基抗体不能阻断结合。虽然VLA - 2整合素促进了CHO细胞中的轮状病毒感染，但很明显仅VLA - 2整合素并不负责轮状病毒与细胞的附着和进入。因此，VLA - 2不参与轮状病毒与细胞的初始附着，但可能在附着后阶段发挥作用。
