title: The Human Cytomegalovirus IE1 Protein Antagonizes PML Nuclear Body-Mediated Intrinsic Immunity via the Inhibition of PML De Novo SUMOylation.
abstract: PML nuclear bodies (NBs) are accumulations of cellular proteins embedded in a scaffold-like structure built by SUMO-modified PML/TRIM19. PML and other NB proteins act as cellular restriction factors against human cytomegalovirus (HCMV); however, this intrinsic defense is counteracted by the immediate early protein 1 (IE1) of HCMV. IE1 directly interacts with the PML coiled-coil domain via its globular core region and disrupts NB foci by inducing a loss of PML SUMOylation. Here, we demonstrate that IE1 acts via abrogating the de novo SUMOylation of PML. In order to overcome reversible SUMOylation dynamics, we made use of a cell-based assay that combines inducible IE1 expression with a SUMO mutant resistant to SUMO proteases. Interestingly, we observed that IE1 expression did not affect preSUMOylated PML; however, it clearly prevented de novo SUMO conjugation. Consistent results were obtained by in vitro SUMOylation assays, demonstrating that IE1 alone is sufficient for this effect. Furthermore, IE1 acts in a selective manner, since K160 was identified as the main target lysine. This is strengthened by the fact that IE1 also prevents As2O3-mediated hyperSUMOylation of K160, thereby blocking PML degradation. Since IE1 did not interfere with coiled-coil-mediated PML dimerization, we propose that IE1 affects PML autoSUMOylation either by directly abrogating PML E3 ligase function or by preventing access to SUMO sites. Thus, our data suggest a novel mechanism for how a viral protein counteracts a cellular restriction factor by selectively preventing the de novo SUMOylation at specific lysine residues without affecting global protein SUMOylation.
  
  
  
  The human cytomegalovirus IE1 protein acts as an important antagonist of a cellular restriction mechanism that is mediated by subnuclear structures termed PML nuclear bodies. This function of IE1 is required for efficient viral replication and thus constitutes a potential target for antiviral strategies. In this paper, we further elucidate the molecular mechanism for how IE1 antagonizes PML NBs. We show that tight binding of IE1 to PML interferes with the de novo SUMOylation of a distinct lysine residue that is also the target of stress-mediated hyperSUMOylation of PML. This is of importance since it represents a novel mechanism used by a viral antagonist of intrinsic immunity. Furthermore, it highlights the possibility of developing small molecules that specifically abrogate this PML-antagonistic activity of IE1 and thus inhibit viral replication.
authors.affiliation: Institute of Clinical and Molecular Virology, Friedrich-Alexander-Universität Erlangen-Nürnberg, Erlangen, Germany.
authors.forename: Eva-Maria
authors.initials: EM
authors.lastname: Schilling
authors.affiliation: Institute of Clinical and Molecular Virology, Friedrich-Alexander-Universität Erlangen-Nürnberg, Erlangen, Germany.
authors.forename: Myriam
authors.initials: M
authors.lastname: Scherer
authors.affiliation: Institute of Clinical and Molecular Virology, Friedrich-Alexander-Universität Erlangen-Nürnberg, Erlangen, Germany.
authors.forename: Nina
authors.initials: N
authors.lastname: Reuter
authors.affiliation: Division of Biotechnology, Friedrich-Alexander-Universität Erlangen-Nürnberg, Erlangen, Germany.
authors.forename: Johannes
authors.initials: J
authors.lastname: Schweininger
authors.affiliation: Division of Biotechnology, Friedrich-Alexander-Universität Erlangen-Nürnberg, Erlangen, Germany.
authors.forename: Yves A
authors.initials: YA
authors.lastname: Muller
authors.affiliation: Institute of Clinical and Molecular Virology, Friedrich-Alexander-Universität Erlangen-Nürnberg, Erlangen, Germany thomas.stamminger@viro.med.uni-erlangen.de.
authors.forename: Thomas
authors.initials: T
authors.lastname: Stamminger
mesh_terms: D002460:Cell Line; D003587:Cytomegalovirus / Q000502:physiology; D003586:Cytomegalovirus Infections / Q000276:immunology / Q000378:metabolism / Q000821:virology; D004795:Enzyme Stability; D006801:Humans; D017874:Immediate-Early Proteins / Q000378:metabolism*; D007109:Immunity; D045586:Intranuclear Inclusion Bodies / Q000378:metabolism*; D009154:Mutation; D000071156:Promyelocytic Leukemia Protein / Q000378:metabolism*; D025841:Small Ubiquitin-Related Modifier Proteins / Q000235:genetics / Q000378:metabolism; D058207:Sumoylation
chemical_list: C088645:IE1 protein, cytomegalovirus; D017874:Immediate-Early Proteins; D000071156:Promyelocytic Leukemia Protein; D025841:Small Ubiquitin-Related Modifier Proteins
keywords: PML nuclear bodies; human cytomegalovirus; immediate early 1; intrinsic immunity; nuclear domain 10; sumoylation