Variation in adenovirus receptor expression and adenovirus vector-mediated transgene expression at defined stages of the cell cycle.
Variation in adenovirus receptor expression and adenovirus vector-mediated transgene expression at defined stages of the cell cycle.
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DOI：
10.1006/mthe.2001.0414
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发表时间：
2001-07
期刊：
影响因子：
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通讯作者：
M. Seidman;Suzanne M. Hogan;Rebecca L. Wendland;S. Worgall;R. Crystal;P. Leopold
中科院分区：
文献类型：
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作者：
M. Seidman;Suzanne M. Hogan;Rebecca L. Wendland;S. Worgall;R. Crystal;P. Leopold
目前已对重组腺病毒（Ad）基因转移载体的感染途径进行了详细的研究，但很少关注细胞生理学对Ad感染结果的影响。基于观察到的Ad感染的克隆细胞群体显示细胞与细胞的衣壳结合的程度的变化，我们假设，细胞周期可能会影响Ad感染的结果。为了解决这一假设，我们评估了Ad与非同步化和非同步化细胞群体中的细胞的关联。在不同步的细胞群体中，Ad与细胞的关联性升高与细胞周期蛋白B1的表达相关，细胞周期蛋白B1是进入有丝分裂M期的标志物。对在M期（使用紫杉醇或诺考达唑）或在S期（使用阿非迪霉素）同步化的细胞群体进行的相同分析证实，与主要处于G（1）和G（2）期的非同步化细胞相比，M期细胞结合3至6倍的衣壳。感染后24小时，载体与细胞的结合增加，转基因表达增加2.5至4倍。对Ad受体的细胞表面表达的评估表明，Ad纤维蛋白的高亲和力柯萨奇-腺病毒受体和Ad五邻体基础蛋白的低亲和力α（v）整联蛋白受体在M期显示出增加的细胞表面表达（分别增加1.5倍和2- 3倍）。这些数据表明，细胞的同质群体的Ad感染可以根据细胞周期阶段而变化，其中增强的Ad结合和表达与M期期间Ad受体的增强表达相关。这些观察结果与了解Ad载体的基因转移机制有关，并有助于设计体内基因转移策略。
Detailed investigations have addressed the infection pathway of recombinant adenovirus (Ad) gene transfer vectors, but little attention has been paid to the influence of cell physiology on the outcome of Ad infection. Based on observations that Ad infection of clonal cell populations show cell-to-cell variability in the extent of capsid binding, we hypothesized that the cell cycle may influence the outcome of Ad infection. To address this hypothesis, we evaluated Ad association with cells in both unsynchronized and pharmacologically synchronized cell populations. In unsynchronized cell populations, elevated Ad association with cells correlated with expression of cyclin B1, a marker of entry into the M phase of mitosis. The same analysis conducted on cell populations that were synchronized at M phase (using paclitaxel or nocodazole) or at S phase (using aphidicolin) confirmed that M phase cells bound three- to sixfold more capsid compared with unsynchronized cells, which are primarily in the G(1) and G(2) phases. The elevated association of vectors with cells translated into 2.5- to 4-fold greater transgene expression 24 hours after infection. Assessment of cell surface expression of Ad receptors demonstrated that both the high-affinity coxsackie-adenovirus receptor for Ad fiber protein and the low-affinity alpha(v) integrin receptor for Ad penton base protein showed increased cell surface expression at M phase (1.5-fold and 2- to 3-fold increases, respectively). These data demonstrate that Ad infection of a homogenous population of cells can vary depending on the cell cycle stage, with enhanced Ad binding and expression correlating with the enhanced expression of Ad receptors during M phase. These observations have relevance to understanding the mechanisms of gene transfer by Ad vectors and should help in the design of in vivo gene transfer strategies.