title: Mutual Interplay between the Human Cytomegalovirus Terminase Subunits pUL51, pUL56, and pUL89 Promotes Terminase Complex Formation.
abstract: Human cytomegalovirus (HCMV) genome encapsidation requires several essential viral proteins, among them pUL56, pUL89, and the recently described pUL51, which constitute the viral terminase. To gain insight into terminase complex assembly, we investigated interactions between the individual subunits. For analysis in the viral context, HCMV bacterial artificial chromosomes carrying deletions in the open reading frames encoding the terminase proteins were used. These experiments were complemented by transient-transfection assays with plasmids expressing the terminase components. We found that if one terminase protein was missing, the levels of the other terminase proteins were markedly diminished, which could be overcome by proteasome inhibition or providing the missing subunit intransThese data imply that sequestration of the individual subunits within the terminase complex protects them from proteasomal turnover. The finding that efficient interactions among the terminase proteins occurred only when all three were present together is reminiscent of a folding-upon-binding principle leading to cooperative stability. Furthermore, whereas pUL56 was translocated into the nucleus on its own, correct nuclear localization of pUL51 and pUL89 again required all three terminase constituents. Altogether, these features point to a model of the HCMV terminase as a multiprotein complex in which the three players regulate each other concerning stability, subcellular localization, and assembly into the functional tripartite holoenzyme.IMPORTANCEHCMV is a major risk factor in immunocompromised individuals, and congenital CMV infection is the leading viral cause for long-term sequelae, including deafness and mental retardation. The current treatment of CMV disease is based on drugs sharing the same mechanism, namely, inhibiting viral DNA replication, and often results in adverse side effects and the appearance of resistant virus strains. Recently, the HCMV terminase has emerged as an auspicious target for novel antiviral drugs. A new drug candidate inhibiting the HCMV terminase, Letermovir, displayed excellent potency in clinical trials; however, its precise mode of action is not understood yet. Here, we describe the mutual dependence of the HCMV terminase constituents for their assembly into a functional terminase complex. Besides providing new basic insights into terminase formation, these results will be valuable when studying the mechanism of action for drugs targeting the HCMV terminase and developing additional substances interfering with viral genome encapsidation.
authors.affiliation: Institute of Virology, Hannover Medical School, Hannover, Germany.
authors.forename: Sebastian
authors.initials: S
authors.lastname: Neuber
authors.affiliation: Institute of Virology, Hannover Medical School, Hannover, Germany.
authors.forename: Karen
authors.initials: K
authors.lastname: Wagner
authors.affiliation: AiCuris Anti-infective Cures GmbH, Wuppertal, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Goldner
authors.affiliation: AiCuris Anti-infective Cures GmbH, Wuppertal, Germany.
authors.forename: Peter
authors.initials: P
authors.lastname: Lischka
authors.affiliation: Institute of Virology, Hannover Medical School, Hannover, Germany.
authors.forename: Lars
authors.initials: L
authors.lastname: Steinbrueck
authors.affiliation: Institute of Virology, Hannover Medical School, Hannover, Germany messerle.martin@mh-hannover.de borst.eva@mh-hannover.de.|German Center for Infection Research (DZIF), Partner Site Hannover-Braunschweig, Hannover-Braunschweig, Germany.
authors.forename: Martin
authors.identifier: http://orcid.org/0000-0002-1227-3933
authors.initials: M
authors.lastname: Messerle
authors.affiliation: Institute of Virology, Hannover Medical School, Hannover, Germany messerle.martin@mh-hannover.de borst.eva@mh-hannover.de.
authors.forename: Eva Maria
authors.initials: EM
authors.lastname: Borst
mesh_terms: D021581:Active Transport, Cell Nucleus / Q000235:genetics; D002460:Cell Line; D022202:Chromosomes, Artificial, Bacterial / Q000235:genetics; D003587:Cytomegalovirus / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D004279:DNA, Viral; D004706:Endodeoxyribonucleases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005347:Fibroblasts / Q000821:virology; D016679:Genome, Viral; D006367:HeLa Cells; D006801:Humans; D046988:Proteasome Endopeptidase Complex / Q000235:genetics; D055550:Protein Stability; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*; D015678:Viral Structural Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D004279:DNA, Viral; C090238:UL56 protein, cytomegalovirus; C436448:UL89 protein, Cytomegalovirus; D014764:Viral Proteins; D015678:Viral Structural Proteins; D004706:Endodeoxyribonucleases; C030894:terminase; D046988:Proteasome Endopeptidase Complex
keywords: complex assembly; cytomegalovirus; nuclear import; protein interactions; protein stability; terminase