The peptidyl-prolyl cis/trans isomerase Pin1 interacts with three early regulatory proteins of human cytomegalovirus

Sonntag

Svrlanska

et al. 2021

Viruses

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Nuclear egress is a common herpesviral process regulating nucleocytoplasmic capsid release. For human cytomegalovirus (HCMV), the nuclear egress complex (NEC) is determined by the pUL50-pUL53 core that regulates multicomponent assembly with NEC-associated proteins and capsids. Recently, NEC crystal structures were resolved for α-, β- and γ-herpesviruses, revealing profound structural conservation, which was not mirrored, however, by primary sequence and binding properties. The NEC binding principle is based on hook-into-groove interaction through an N-terminal hook-like pUL53 protrusion that embraces an α-helical pUL50 binding groove. So far, pUL50 has been considered as the major kinase-interacting determinant and massive phosphorylation of pUL50-pUL53 was assigned to NEC formation and functionality. Here, we addressed the question of phenotypical changes of ORF-UL50-mutated HCMVs. Surprisingly, our analyses did not detect a predominant replication defect for most of these viral mutants, concerning parameters of replication kinetics (qPCR), viral protein production (Western blot/CoIP) and capsid egress (confocal imaging/EM). Specifically, only the ORF-UL50 deletion rescue virus showed a block of genome synthesis during late stages of infection, whereas all phosphosite mutants exhibited marginal differences compared to wild-type or revertants. These results (i) emphasize a rate-limiting function of pUL50 for nuclear egress, and (ii) demonstrate that mutations in all mapped pUL50 phosphosites may be largely compensated. A refined mechanistic concept points to a multifaceted nuclear egress regulation, for which the dependence on the expression and phosphorylation of pUL50 is discussed.

Section: Patterns Of Expression and Protein-protein Interaction Of Orf-ul50 Deletion And Phosphosite Mutantssupporting

confidence: 89%

Phenotypical Characterization of the Nuclear Egress of Recombinant Cytomegaloviruses Reveals Defective Replication upon ORF-UL50 Deletion but Not pUL50 Phosphosite Mutation

Sonntag

Svrlanska

et al. 2021

Viruses

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“…Functional replacement between herpesviral kinases has been demonstrated with specific examples [ 89 , 90 , 91 , 92 ]. Consequently, site-specific Ser22-phosphorylated lamins are recognized by the cellular peptidyl-prolyl cis/trans isomerase Pin1, which isomerizes the lamins towards conformational reorganization and, thus, disassembles the lamina network to provide the capsids access to the INM [ 26 , 41 , 93 ]. Concerning the membrane-specific effects of the nuclear egress pathway, several highly valuable model systems have been established, in particular representing α-herpesvirus infections [ 6 , 7 , 94 , 95 , 96 ].…”

Section: Main Functionality and Regulatory Roles Shared By Herpesvira...mentioning

confidence: 99%

‘Come together’—The Regulatory Interaction of Herpesviral Nuclear Egress Proteins Comprises Both Essential and Accessory Functions

Marschall

2022

Cells

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Herpesviral nuclear egress is a fine-tuned regulatory process that defines the nucleocytoplasmic release of viral capsids. Nuclear capsids are unable to traverse via nuclear pores due to the fact of their large size; therefore, herpesviruses evolved to develop a vesicular transport pathway mediating the transition across the two leaflets of the nuclear membrane. The entire process involves a number of regulatory proteins, which support the local distortion of the nuclear envelope. In the case of the prototype species of β-Herpesvirinae, the human cytomegalovirus (HCMV), the nuclear egress complex (NEC) is determined by the core proteins pUL50 and pUL53 that oligomerize, form capsid docking lattices and mediate multicomponent assembly with NEC-associated viral and cellular proteins. The NEC-binding principle is based on the hook-into-groove interaction through an N-terminal hook-like pUL53 protrusion that embraces an α-helical pUL50 binding groove. Thus far, the function and characteristics of herpesviral core NECs have been well studied and point to the groove proteins, such as pUL50, as the multi-interacting, major determinants of NEC formation and egress. This review provides closer insight into (i) sequence and structure conservation of herpesviral core NEC proteins, (ii) experimentation on cross-viral core NEC interactions, (iii) the essential functional roles of hook and groove proteins for viral replication, (iv) an establishment of assay systems for NEC-directed antiviral research and (v) the validation of NEC as putative antiviral drug targets. Finally, this article provides new insights into the conservation, function and antiviral targeting of herpesviral core NEC proteins and, into the complex regulatory role of hook and groove proteins during the assembly, egress and maturation of infectious virus.

“…To illustrate this process, the strategies of pharmacological Pin1 inhibition or genetic Pin1 knockout provided conclusive evidence that Pin1 promotes the disassembly of the nuclear lamina during HCMV replication [ 10 , 12 ]. Very recently, we were able to provide additional evidence for the relevance of Pin1 in HCMV replication, in that data derived from several independent approaches demonstrated an interaction of Pin1 with the viral proteins pUL50, pUL69 and pUL44 [ 75 , 76 ]. Whether the regulatory impact of Pin1, especially its lamin-directed role in nuclear egress, is specific for HCMV, or whether Pin1 activity is likewise a regulatory cofactor for other α-, β- and γ-herpesviruses has still to be clarified.…”

Section: Specific Functional Properties Of Necs and Egress Processmentioning

confidence: 99%

Nuclear Egress Complexes of HCMV and Other Herpesviruses: Solving the Puzzle of Sequence Coevolution, Conserved Structures and Subfamily-Spanning Binding Properties

Marschall

Conrad

et al. 2020

Viruses

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Herpesviruses uniquely express two essential nuclear egress-regulating proteins forming a heterodimeric nuclear egress complex (core NEC). These core NECs serve as hexameric lattice-structured platforms for capsid docking and recruit viral and cellular NEC-associated factors that jointly exert nuclear lamina as well as membrane-rearranging functions (multicomponent NEC). The regulation of nuclear egress has been profoundly analyzed for murine and human cytomegaloviruses (CMVs) on a mechanistic basis, followed by the description of core NEC crystal structures, first for HCMV, then HSV-1, PRV and EBV. Interestingly, the highly conserved structural domains of these proteins stand in contrast to a very limited sequence conservation of the key amino acids within core NEC-binding interfaces. Even more surprising, although a high functional consistency was found when regarding the basic role of NECs in nuclear egress, a clear specification was identified regarding the limited, subfamily-spanning binding properties of core NEC pairs and NEC multicomponent proteins. This review summarizes the evolving picture of the relationship between sequence coevolution, structural conservation and properties of NEC interaction, comparing HCMV to α-, β- and γ-herpesviruses. Since NECs represent substantially important elements of herpesviral replication that are considered as drug-accessible targets, their putative translational use for antiviral strategies is discussed.