UL28 and UL33 homologs of Marek\u2019s disease virus terminase complex involved in the regulation of cleavage and packaging of viral DNA are indispensable for replication in cultured cells

Aijun Sun,Yanan Wu,Shuaikang Yang,Man Teng,Rui Wang,Jun Luo,Venugopal Nair,Yijie Xu,Luping Zheng,Yongxiu Yao,Guoqing Zhuang,Xiaojing Zhu,Gaiping Zhang

doi:10.1186/s13567-021-00901-5

Abstract

Processing and packaging of herpesvirus genomic DNA is regulated by a packaging-associated terminase complex comprising of viral proteins pUL15, pUL28 and pUL33. Marek’s disease virus (MDV) homologs UL28 and UL33 showed conserved functional features with high sequence identity with the corresponding Herpes simplex virus 1 (HSV-1) homologs. As part of the investigations into the role of the UL28 and UL33 homologs of oncogenic MDV for DNA packaging and replication in cultured cells, we generated MDV mutant clones deficient in UL28 or UL33 of full-length MDV genomes. Transfection of UL28- or UL33-deleted BAC DNA into chicken embryo fibroblast (CEF) did not result either in the production of visible virus plaques, or detectable single cell infection after passaging onto fresh CEF cells. However, typical MDV plaques were detectable in CEF transfected with the DNA of revertant mutants where the deleted genes were precisely reinserted. Moreover, the replication defect of the UL28-deficient mutant was completely restored when fragment encoding the full UL28 gene was co-transfected into CEF cells. Viruses recovered from the revertant construct, as well as by the UL28 co-transfection, showed replication ability comparable with parental virus. Furthermore, the transmission electron microscopy study indicated that immature capsids were assembled without the UL28 expression, but with the loss of infectivity. Importantly, predicted three-dimensional structures of UL28 between MDV and HSV-1 suggests conserved function in virus replication. For the first time, these results revealed that both UL28 and UL33 are essential for MDV replication through regulating DNA cleavage and packaging.

Highlights

Herpesviruses are categorized to α, β and γ subfamilies based on genome structure and biology [1]
During herpesvirus replication, an assembled icosahedral capsid was initially formed, and viral genomic DNA was packaged into through the entrance and exit pore of portal vertex, which is driven by terminase complex [12]
We have predicted and conducted experiments to identify the role of the UL28 and UL33 in Marek’s disease virus (MDV) replication in vitro

Summary

Introduction

Herpesviruses are categorized to α, β and γ subfamilies based on genome structure and biology [1]. Genome alignment analysis revealed the high conservation of MDV serotypes especially in UL and US regions, which encodes highly conserved virus replication related elements, such as glycoproteins, tegument proteins and capsid-associated proteins [5, 6]. MDVencoded tegument proteins perform both regulatory and structural roles in the viral replication. One of the major tegument protein VP22, encoded by UL49, was indispensable for the propagation of MDV mainly relying on its N-terminus central domain, which might affect viral replication through modulating cell cycle metabolism [10]. While UL47 was found to be expressed at very low level in infected cultured cells, UL46, UL48 and UL49 showed enhanced expression suggesting a potential regulatory role in viral replication [11]

Methods

Results

Conclusion