Role of the serine-threonine kinase PAK-1 in myxoma virus replication
Role of the serine-threonine kinase PAK-1 in myxoma virus replication
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DOI：
10.1128/jvi.77.10.5877-5888.2003
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发表时间：
2003-05-01
影响因子：
5.4
通讯作者：
McFadden, G
中科院分区：
文献类型：
--
作者：
Johnston, JB;Barrett, JW;McFadden, G
颠覆或占用细胞信号转导通路是病毒用来促进受感染细胞内支持病毒复制周期的环境的一种常见策略。利用3T3小鼠成纤维细胞亚群，我们研究了宿主丝氨酸-苏氨酸激酶（STKs）作为允许表型的潜在介质的作用。这些亚群在支持黏液瘤病毒（MV）复制能力方面存在差异。允许和非允许的3T3细胞都支持相同水平的病毒粒子结合、进入和早期病毒基因表达，这表明3T3细胞中的MV趋向性不是由受体介导的进入决定的。相反，在限制性3T3细胞中，晚期病毒基因表达和病毒DNA复制被选择性地破坏。添加特异性蛋白激酶抑制剂，其中许多具有影响STKs p21激活的激酶1 （PAK-1）和Raf-1活性的能力，可减弱W在允许的3T3细胞中的复制。Western blot检测磷酸化形式的PAK-1 （Thr423）和Raf-1 （Ser338）证实，在W感染或γ干扰素治疗后，这些激酶在允许细胞中被激活，但在限制性3T3细胞中，这两种激酶的激活形式大大减少或不存在。这些激活的生物学意义通过使用PAK-1的自抑制结构域（氨基酸83至149）来证明，其表达降低了允许的3T3细胞中MV感染的效率，同时降低了PAK-1的激活。相比之下，在限制性3T3细胞中，过表达构成活性的PAK-1 （T423E）突变体增加了MV复制。这些观察结果表明，通过细胞STKs诱导的信号传导可能在决定宿主细胞对痘病毒感染的容错性方面发挥重要作用。
Subversion or appropriation of cellular signal transduction pathways is a common strategy employed by viruses to promote an environment within infected cells that supports the viral replicative cycle. Using subsets of 3T3 murine fibroblasts previously shown to differ in their ability to support myxoma virus (MV) replication, we investigated the role of host serine-threonine kinases (STKs) as potential mediators of the permissive phenotype. Both permissive and nonpermissive 3T3 cells supported equivalent levels of virion binding, entry, and early virus gene expression, indicating that MV tropism in 3T3 cells was not determined by receptormediated entry. In contrast, late virus gene expression and viral DNA replication were selectively compromised in restrictive 3T3 cells. Addition of specific protein kinase inhibitors, many of which shared the ability to influence the activity of the STKs p21-activated kinase 1 (PAK-1) and Raf-1 attenuated W replication in permissive 3T3 cells. Western blot detection of the phosphorylated forms of PAK-1 (Thr423) and Raf-1 (Ser338) confirmed activation of these kinases in permissive cells after W infection or gamma interferon treatment, but the activated forms of both kinases were greatly reduced or absent in restrictive 3T3 cells. The biological significance of these activations was demonstrated by using the autoinhibitory domain of PAK-1 (amino acids 83 to 149), expression of which reduced the efficiency of MV infection in permissive 3T3 cells concurrent with a decrease in PAK-1 activation. In comparison, overexpression of a constitutively active PAK-1 (T423E) mutant increased MV replication in restrictive 3T3 cells. These observations suggest that induced signaling via cellular STKs may play important roles in determining the permissiveness of host cells to poxvirus infection.