Mutations in the adenovirus-encoded single-stranded DNA binding protein that result in altered accumulation of early and late viral RNAs.
Mutations in the adenovirus-encoded single-stranded DNA binding protein that result in altered accumulation of early and late viral RNAs.
复制标题
腺病毒编码的单链 DNA 结合蛋白发生突变，导致早期和晚期病毒 RNA 的积累发生改变。
DOI：
10.1006/viro.1995.1457
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发表时间：
1995
期刊：
影响因子：
3.7
通讯作者：
Kitchingman,GR
中科院分区：
文献类型：
--
作者：
Kitchingman,GR
腺病毒编码一个72 kDa的单链DNA结合蛋白（DBP），它是病毒DNA复制所必需的，并参与控制病毒RNA代谢。对DBP的温度敏感性（ts）和定点突变体的研究已经确定了至少四个参与结合单链DNA并因此参与DNA复制的蛋白质区域。其中两个ts突变体，Ad 2 ts 111 A和Ad 2ND 1 + ts 23，在DNA结合和支持体外DNA复制方面有缺陷。它们对病毒RNA代谢的影响在这里介绍。在感染的早期和晚期，相对于野生型病毒，在Ad 2 ts 111 A-和Ad 2ND 1 + ts 23-感染的细胞中，来自病毒E3区的RNA的积累增加至7倍。这种效应是温度无关的，似乎涉及核RNA的稳定性。在野生型病毒感染的细胞中，病毒E1 B和E4 RNA的稳态水平在病毒DNA复制开始后增加，但在ts 111 A或ts 23感染的细胞中则没有。E1 B和E4 RNA在感染后期的增加是由于mRNA的稳定。在感染ts 23和ts 111 A的细胞中，L3和L5 RNA的稳态水平比野生型病毒高2 - 4倍。这些差异都没有观察到感染后的细胞与温度无关的回复突变的ts 111，表明DBP的突变负责的表型。然而，对于E3效应，DBP突变引起的变化似乎不是蛋白质正常功能变化的结果，因为基本上DBP阴性的病毒（Ad 5dl 802）在感染早期与野生型病毒相比在E3 RNA代谢方面没有差异。这些结果表明DBP的氨基酸280和282的突变显著干扰病毒RNA代谢的正常调节。这些效应明显不同于在其他早期基因中含有突变的腺病毒中观察到的表型，也不同于其他人归因于ts 125突变的表型。这些结果进行了讨论的机制，早期和晚期病毒RNA代谢的控制和DBP突变对他们的可能影响。
Adenovirus encodes a 72-kDa single-stranded DNA binding protein (DBP) that is necessary for viral DNA replication and is involved in controlling viral RNA metabolism. Studies of temperature-sensitive (ts) and site-directed mutants of the DBP have identified at least four regions of the protein involved in binding to single-stranded DNA and hence in DNA replication. Two of the ts mutants, Ad2ts111A and Ad2ND1+ts23, are deficient in DNA binding and in supporting in vitro DNA replication. Their effects on viral RNA metabolism are presented here. At early and late times of infection, accumulation of RNAs from the viral E3 region is increased up to sevenfold in Ad2ts111A- and Ad2ND1+ts23-infected cells relative to wild-type virus. This effect is temperature-independent and seems to involve nuclear RNA stability. Steady-state levels of the viral E1B and E4 RNAs increase following the onset of viral DNA replication in cells infected by the wild-type virus, but not in cells infected by ts111A or ts 23. The increase in E1B and E4 RNAs at late times of infection is due to a stabilization of the mRNA. The steady-state levels of L3 and L5 RNAs are two- to fourfold higher in cells infected with ts23 and ts111A than with wild-type virus. None of these differences were observed following infection of cells with a temperature-independent revertant of ts111, indicating that the mutations in the DBP were responsible for the phenotypes. However, for the E3 effect, the change brought about by the mutations in the DBP does not seem to be the result of a change in the normal function of the protein, as an essentially DBP-negative virus (Ad5dl802) shows no differences in E3 RNA metabolism compared to wild-type virus at early times of infection. These results demonstrate that mutation of amino acids 280 and 282 of the DBP significantly perturbs the normal regulation of viral RNA metabolism. These effects clearly differ from the phenotypes observed with adenoviruses containing mutations in other early genes and from those ascribed to the ts125 mutation by others. These results are discussed in terms of the mechanisms by which early and late viral RNA metabolism are controlled and the possible effects of the DBP mutations on them.