A mobile functional region of Kaposi's sarcoma-associated herpesvirus ORF50 protein independently regulates DNA binding and protein abundance
A mobile functional region of Kaposi's sarcoma-associated herpesvirus ORF50 protein independently regulates DNA binding and protein abundance
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DOI：
10.1128/jvi.00862-08
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发表时间：
2008-10-01
影响因子：
5.4
通讯作者：
Miller, George
中科院分区：
文献类型：
--
作者：
Chang, Pey-Jium;Shedd, Duane;Miller, George
卡波西肉瘤相关疱疹病毒（KSHV）开放阅读框50（ORF 50）编码的蛋白质作为转录激活因子发挥作用，并在裂解性病毒DNA复制中介导从潜伏病毒感染到裂解期的转换。在这里，我们确定了ORF 50蛋白的调控区域，独立控制DNA结合和蛋白质的丰度。一个区域含有位于ORF 50的氨基酸（aa）490和535之间的DNA结合抑制序列（DBIS）。该序列中的一簇碱性氨基酸在抑制DNA结合中是重要的。DBIS可以在ORF 50蛋白的N或C末端或内部起作用。由于DBIS在从大肠杆菌纯化的ORF 50蛋白中具有功能，因此它可能通过分子内机制发挥作用。第二个调节区，蛋白丰度调节信号（PARS），由两个组成部分。PARS的组分I与DBIS重叠，但可通过碱性氨基酸残基的特异性取代与DBIS区分。PARS的第二部分位于aa 590和650之间。任一组分的突变或缺失导致ORF 50蛋白的大量表达。当双组分PARS融合到异源蛋白Glucose S-转移酶时，融合蛋白是不稳定的。DBIS或PARS中的突变损害ORF 50激活直接和间接靶病毒启动子的能力。由于这些重叠的调控基序位于C-末端的反式激活结构域，它们可能是重要的控制ORF 50蛋白的许多行动。
The protein encoded by open reading frame 50 (ORF50) of Kaposi's sarcoma-associated herpesvirus (KSHV) functions as a transcriptional activator and in lytic viral DNA replication to mediate the switch from latent viral infection to the lytic phase. Here we identify regulatory regions of ORF50 protein that independently control DNA binding and abundance of the protein. One region contains a DNA-binding inhibitory sequence (DBIS) located between amino acids (aa) 490 and 535 of ORF50. A cluster of basic amino acids in this sequence is important in inhibiting DNA binding. The DBIS can function at the N or C terminus or internally in the ORF50 protein. Since the DBIS is functional in ORF50 protein purified from Escherichia coli, it is likely to work through an intramolecular mechanism. The second regulatory region, a protein abundance regulatory signal (PARS), consists of two components. Component I of the PARS overlaps the DBIS but can be differentiated from the DBIS by specific substitution of basic amino acid residues. Component II of PARS is located between aa 590 and 650. Mutation or deletion of either component results in abundant expression of ORF50 protein. When the two-component PARS was fused to a heterologous protein, Glutathione S-transferase, the fusion protein was unstable. Mutations in the DBIS or PARS impair the capacity of ORF50 to activate direct and indirect target viral promoters. Since these overlapping regulatory motifs are located in the C-terminal transactivation domain, they are likely to be important in controlling many actions of ORF50 protein.