The role of protein kinase A regulation of the E6 PDZ-binding domain during the differentiation-dependent life cycle of human papillomavirus type 18.
The role of protein kinase A regulation of the E6 PDZ-binding domain during the differentiation-dependent life cycle of human papillomavirus type 18.
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DOI：
10.1128/jvi.01234-13
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发表时间：
2013-09
影响因子：
5.4
通讯作者：
Roberts S
中科院分区：
文献类型：
--
作者：
Delury CP;Marsh EK;James CD;Boon SS;Banks L;Knight GL;Roberts S
高危α型人乳头瘤病毒（HPV）E6蛋白通过使用C-末端PDZ结合基序（PBM）靶向一组选定的含PSD 95/DLG 1/ZO 1（PDZ）结构域的蛋白，这是一种可通过蛋白激酶A（PKA）磷酸化E6 PBM负调控的相互作用。在此，我们突变了与HPV 18基因组中的E6 PBM部分重叠的典型PKA识别基序（E6153 PKA），并将该突变对原代角质形成细胞中HPV 18生命周期的影响与野生型基因组和缺乏E6 PBM的第二突变基因组（E6ΔPDZ）进行了比较。与携带野生型基因组的细胞相比，E6的PKA识别的丧失与含基因组的细胞的生长增加相关，并且在分层时，具有更多的增生表型，在上基底层中S期感受态细胞的数量增加，而E6ΔPDZ基因组则相反。此外，含有野生型基因组的细胞的生长对PKA活性的变化敏感，并且这些变化与E6 PBM的磷酸化增加相关。与E6ΔPDZ基因组形成鲜明对比的是，E6153 PKA突变对病毒基因组扩增或晚期蛋白表达没有有害影响。我们的数据表明，E6 PBM功能的差异调节磷酸化在HPV 18的生命周期。我们推测，蛋白激酶信号通路的扰动可能导致E6 PBM功能的变化，这反过来可能与肿瘤的促进和进展有关。
Human papillomavirus (HPV) E6 proteins of high-risk alpha types target a select group of PSD95/DLG1/ZO1 (PDZ) domain-containing proteins by using a C-terminal PDZ-binding motif (PBM), an interaction that can be negatively regulated by phosphorylation of the E6 PBM by protein kinase A (PKA). Here, we have mutated the canonical PKA recognition motif that partially overlaps with the E6 PBM in the HPV18 genome (E6153PKA) and compared the effect of this mutation on the HPVl8 life cycle in primary keratinocytes with the wild-type genome and with a second mutant genome that lacks the E6 PBM (E6ΔPDZ). Loss of PKA recognition of E6 was associated with increased growth of the genome-containing cells relative to cells carrying the wild-type genome, and upon stratification, a more hyperplastic phenotype, with an increase in the number of S-phase competent cells in the upper suprabasal layers, while the opposite was seen with the E6ΔPDZ genome. Moreover, the growth of wild-type genome-containing cells was sensitive to changes in PKA activity, and these changes were associated with increased phosphorylation of the E6 PBM. In marked contrast to E6ΔPDZ genomes, the E6153PKA mutation exhibited no deleterious effects on viral genome amplification or expression of late proteins. Our data suggest that the E6 PBM function is differentially regulated by phosphorylation in the HPV18 life cycle. We speculate that perturbation of protein kinase signaling pathways could lead to changes in E6 PBM function, which in turn could have a bearing on tumor promotion and progression.