The "Bridge" in the Epstein-Barr Virus Alkaline Exonuclease Protein BGLF5 Contributes to Shutoff Activity during Productive Infection
The "Bridge" in the Epstein-Barr Virus Alkaline Exonuclease Protein BGLF5 Contributes to Shutoff Activity during Productive Infection
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DOI：
10.1128/jvi.00309-12
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发表时间：
2012-09-01
影响因子：
5.4
通讯作者：
Ressing, Maaike E.
中科院分区：
文献类型：
--
作者：
Horst, Danielle;Burmeister, Wim P.;Ressing, Maaike E.
人类疱疹病毒EB病毒的复制严重损害细胞蛋白质合成。这种关闭表型是由于早期裂解相蛋白BGLF 5表达后mRNA降解所致。有趣的是，BGLF 5是病毒DNA酶或碱性核酸外切酶，其同系物存在于整个疱疹病毒家族中。在生产性感染过程中，这种DNA酶对于病毒基因组的加工和包装至关重要。与这种广泛保守的DNA酶活性相反，关闭仅由γ疱疹病毒亚科的碱性核酸外切酶介导。在这里，我们表明，BGLF 5可以降解细胞和病毒来源的mRNA，无论多聚腺苷酸化。此外，通过BGLF 5的关闭诱导细胞溶质poly（A）结合蛋白的核重新定位。在最近解析的BGLF 5结构的指导下，产生突变体并分析其对DNA酶和关闭活性的功能后果。一方面，破坏DNA酶活性的点突变也阻断RNA酶功能，这意味着两种活性共享一个催化位点。另一方面，其他突变更具选择性，对DNA降解或关闭具有更显著的影响。后者的结果是指示的一个阿托西肽结合位点，是部分共享的DNA和RNA。为此，跨越BGLF 5的活性位点峡谷的柔性“桥”似乎有助于与RNA底物的相互作用。这些发现扩展了我们对BGLF 5关闭功能的分子基础的理解，BGLF 5在γ疱疹病毒中是保守的，但在α和β疱疹病毒中不是。
Replication of the human herpesvirus Epstein-Barr virus drastically impairs cellular protein synthesis. This shutoff phenotype results from mRNA degradation upon expression of the early lytic-phase protein BGLF5. Interestingly, BGLF5 is the viral DNase, or alkaline exonuclease, homologues of which are present throughout the herpesvirus family. During productive infection, this DNase is essential for processing and packaging of the viral genome. In contrast to this widely conserved DNase activity, shutoff is only mediated by the alkaline exonucleases of the subfamily of gammaherpesviruses. Here, we show that BGLF5 can degrade mRNAs of both cellular and viral origin, irrespective of polyadenylation. Furthermore, shutoff by BGLF5 induces nuclear relocalization of the cytosolic poly(A) binding protein. Guided by the recently resolved BGLF5 structure, mutants were generated and analyzed for functional consequences on DNase and shutoff activities. On the one hand, a point mutation destroying DNase activity also blocks RNase function, implying that both activities share a catalytic site. On the other hand, other mutations are more selective, having a more pronounced effect on either DNA degradation or shutoff. The latter results are indicative of an oligonucleotide-binding site that is partially shared by DNA and RNA. For this, the flexible "bridge" that crosses the activesite canyon of BGLF5 appears to contribute to the interaction with RNA substrates. These findings extend our understanding of the molecular basis for the shutoff function of BGLF5 that is conserved in gammaherpesviruses but not in alpha- and betaherpesviruses.