Wagner M, Michel D, Schaarschmidt P, Vaida B, Jonjic S, Messerle M, Mertens T, Koszinowski U
Max von Pettenkofer Institut für Hygiene und Medizinische Mikrobiologie, Ludwig-Maximilians-Universität München, Munich, Germany.
J Virol. 2000 Nov;74(22):10729-36. doi: 10.1128/jvi.74.22.10729-10736.2000.
The UL97 protein (pUL97) of human cytomegalovirus (HCMV) is a protein kinase that also phosphorylates ganciclovir (GCV), but its biological function is not yet clear. The M97 protein (pM97) of mouse cytomegalovirus (MCMV) is the homolog of pUL97. First, we studied the consequences of genetic replacement of M97 by UL97. Using the infectious bacterial plasmid clone of the full-length MCMV genome (M. Wagner, S. Jonjic, U. H. Koszinowski, and M. Messerle, J. Virol. 73:7056-7060, 1999), we replaced the M97 gene with the UL97 gene and constructed an MCMV M97 deletion mutant and a revertant virus. In addition, pUL97 and pM97 were expressed by recombinant vaccinia virus to compare both for known functions. Remarkably, pM97 proved not to be the reason for the GCV sensitivity of MCMV. When expressed by the recombinant MCMV, however, pUL97 was phosphorylated and endowed MCMV with the capacity to phosphorylate GCV, thereby rendering MCMV more susceptible to GCV. We found that deletion of pM97, although it is not essential for MCMV replication, severely affected virus growth. This growth deficit was only partially amended by pUL97 expression. When expressed by recombinant vaccinia viruses, both proteins were phosphorylated and supported phosphorylation of GCV, but pUL97 was about 10 times more effective than pM97. One hint of the functional differences between the proteins was provided by the finding that pUL97 accumulates in the nucleus, whereas pM97 is predominantly located in the cytoplasm of infected cells. In vivo testing revealed that the UL97-MCMV recombinant should allow evaluation of novel antiviral drugs targeted to the UL97 protein of HCMV in mice.
人巨细胞病毒（HCMV）的UL97蛋白（pUL97）是一种蛋白激酶，它也能磷酸化更昔洛韦（GCV），但其生物学功能尚不清楚。小鼠巨细胞病毒（MCMV）的M97蛋白（pM97）是pUL97的同源物。首先，我们研究了用UL97基因替换M97基因的遗传学后果。利用全长MCMV基因组的感染性细菌质粒克隆（M. Wagner、S. Jonjic、U. H. Koszinowski和M. Messerle，《病毒学杂志》73:7056 - 7060，1999年），我们用UL97基因替换了M97基因，并构建了一个MCMV M97缺失突变体和一个回复病毒。此外，通过重组痘苗病毒表达pUL97和pM97以比较两者的已知功能。值得注意的是，pM97并非MCMV对GCV敏感的原因。然而，当由重组MCMV表达时，pUL97被磷酸化并赋予MCMV磷酸化GCV的能力，从而使MCMV对GCV更敏感。我们发现，pM97的缺失虽然对MCMV复制不是必需的，但严重影响病毒生长。这种生长缺陷仅部分被pUL97的表达所改善。当由重组痘苗病毒表达时，两种蛋白都被磷酸化并支持GCV的磷酸化，但pUL97的效果比pM97高约10倍。pUL97在细胞核中积累，而pM97主要位于感染细胞的细胞质中，这一发现提示了这两种蛋白之间的功能差异。体内试验表明，UL97 - MCMV重组体应有助于在小鼠中评估针对HCMV的UL97蛋白的新型抗病毒药物。
